Mardiyyah/TAPT_data_V2_split · Datasets at Hugging Face

PMCID stringclasses 24 values	Title stringclasses 24 values	Sentences stringlengths 2 40.7k
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	The hallmarks of diabetes-induced RN, which include neural-cell apoptosis, diminished retinal neuronal function, and reactive gliosis, have been observed to occur prior to the onset of overt microangiopathy in experimental models of DR, diabetic patients, and post-mortem human retinas .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Müller Glia cells (MGs) are the main retinal glial cells, extending throughout the entire thickness from the inner to the outer limiting membranes, providing an anatomical link between the retinal neurons and the retinal blood vessels and thus responsible for the homeostatic and metabolic support of retinal nerve cells .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	MGs also play a pivotal role in maintaining neuronal health through the recycling of neurotransmitters and the prevention of neurotoxicity resulting from glutamate excess.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Additionally, they regulate ionic balance by buffering K ions and supply lactate as an energy source for photoreceptors .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In response to diabetic stress, MGs undergo morphological changes and exhibit an increased expression of the intermediate filament proteins, such as glial fibrillary acidic protein (GFAP), a key marker of reactive gliosis .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Reactive gliosis plays a dual role in the context of retinal damage, with the potential to both protect against damage and contribute to its progression in a diabetic milieu.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	The formation of a glial scar, while stabilizing the tissue structure, may also impair neuronal function through the release of proinflammatory mediators .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	The ambivalent role of MGs gliosis in DR underscores the complexity of their involvement, thereby emphasizing the necessity for a comprehensive understanding of the regulatory mechanisms involved.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	The impact of fluctuating glycemic conditions on MGs responses introduces an additional layer of complexity.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Fluctuations in blood glucose levels have been demonstrated to trigger diverse signaling pathways within MGs, resulting in an increase of the inner nuclear layer (INL) in subjects with diabetes , potentially leading to differential activation states or pathological outcomes .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	As a matter of fact, in recent years significant advancements have been made in understanding the pathophysiological roles of MGs in DR .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Studies in lower vertebrates, including fish and amphibians, have demonstrated that, following reactive gliosis, the sonic hedgehog (SHH) signaling pathway is activated, thereby promoting the reprogramming of MGs into various retinal cell types and contributing to retinal regeneration .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	The capacity for retinal regeneration is notably limited in mammals, including humans.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In response to injury, MGs typically undergo a process of reactive gliosis, which often results in scarring rather than regeneration .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	To achieve significant regenerative processes, additional manipulations of the SHH pathway may be required .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Recent studies have identified the potential genes involved in reprogramming MGs into retinal neurons, such as Ascl1, Sox2, and Lin28, which can be modulated to stimulate functional neuronal regeneration .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Despite the advancement of knowledge regarding the neurogenic process, however, the existing literature lacks insights into the influence of glucose metabolism on these processes.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In particular, as far as we know, there is no evidence in the literature indicating the effect of glucose metabolism on retinal neurogenesis.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Our previous in vitro studies, involving rat retinal Müller cell line rMC-1, have demonstrated that these cells are activated in response to both high glucose levels and glucose fluctuations (GFs), and that this activation is associated with cellular increased levels of GFAP and aquaporin-4 (AQPs) .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	These findings contributed to elucidating the dynamic responses of Müller cells under varied glycaemic conditions, suggesting the existence of a complex regulatory mechanism sensitive to metabolic fluctuations.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	However, to better understand the mechanisms underlying DR in humans, it is essential to investigate whether or not similar responses occur in human Müller cell lines, as cellular responses may differ between animal models and humans, particularly due to differences in complexity and disease progression.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	The present study aims to fill this gap by investigating the gliotic response of human Müller cell line MIO-M1, a well-established model in human retinal research, exposed to sustained high-glucose and glucose fluctuation treatments that mimic the different glycemic conditions that can occur in diabetic patients.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Furthermore, the study aims to investigate the reprogramming potential of MIO-M1 cells under different glucose stress conditions, focusing on their potential for dedifferentiation and neurogenesis.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	These processes are of significant interest in understanding how glial cells respond to pathological conditions such as DR.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	This approach allowed one to explore not only the gliotic response but also the potential for reprogramming Müller cells into a more progenitor-like state, a key aspect for the development of future regenerative therapies.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	The results obtained might contribute to elucidating the dynamic responses of MGs under different glycemic conditions, suggesting the existence of a complex regulatory mechanism sensitive to glucose metabolic stress.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	WB analysis carried out on MIO-M1 cells cultured under HG condition showed a significant increase in GFAP levels compared to MIO-M1 cells cultured under NG condition (fold change of about 2 in HG vs. NG), indicating an enhanced gliotic response under hyperglycemic stress conditions (Figure 1A,B).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Such significant increases of GFAP levels was also observed when NG MIO-M1 cells were exposed to sustained high-glucose (II) and GF (III) treatments compared to their basal glucose (I) (Figure 2A,B and Figure S1).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Although a clear trend towards increased GFAP expression was also observed across GF treatments IV and V, these increases were not statistically significant (Figure 2A,B).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In contrast, MIO-M1 cells cultured under HG condition showed a markedly different response, with no significant changes in GFAP levels when these cells were exposed to sustained high-glucose (II) and GF (III–V) treatments compared to their basal glucose treatment (I) (Figure 2A,B).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	The impact of the experimental culture conditions described above on GFAP expression in MIO-MI cells was also investigated by IF, and cellular bipolar or radial morphologies, typical of quiescent and activated Müller cells, respectively , were also recorded.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Under NG condition, GFAP expression was observed in a limited number of MIO-MI cells, approximately 15%, which were GFAP positive, consistent with literature data .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Typical of low active Müller cell status, GFAP staining in NG MIO-M1 cells was faint, while bipolar or radial morphologies were equally distributed (Figure 3A,B).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In contrast, MIO-M1 cells maintained in the HG condition showed intense GFAP staining, with almost 80% displaying radial morphology (Figure 3A,B), indicating that these cells are in a more activated state.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Similarly to the WB results, in NG MIO-M1cells, subjected to all different glucose treatments (II–V), a significant increase in the GFAP intensity and radial morphology were observed, compared to the basal glucose treatment (I) (NG: 47.85% condition I vs. 69,63% condition II; 77.80% condition III; 74.21% condition IV; 81.42% condition V) (Figure 4A,B).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In contrast, in HG MIO-M1 cells subjected to any experimental glucose treatments, no significant change in GFAP positivity or intensity and morphology were detected compared to the basal glucose treatment (Figure 4A,C).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	IF for vimentin filaments, another well-established marker of Müller cell stress response, on MIO-M1 cells subjected to all different glucose treatments gave similar results to those described for GFAP and cell morphology changes.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Briefly, NG MIO-M1 cells in the basal glucose treatment (I) exhibited intact cytoskeletal integrity, with well-organized and linear vimentin filaments.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Following the exposure of NG MIO-M1 cells to sustained high-glucose (II) and GF (III–V) treatments, a significant increase in vimentin expression was observed, compared to the basal glucose treatment (I).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	This increase was associated with a more disorganized appearance of vimentin filaments, reflecting increased cellular stress and cytoskeletal reorganization (Figure 5).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Such a result was also observed in HG MIO-M1 cells maintained in the basal glucose treatment (I) and when these cells were exposed to all different glucose treatments (II–V) (Figure 5).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In order to investigate the activation of the neurogenic potential of MIO-M1 cells under the various glucose treatments investigated here, the expression and cellular localization of sonic hedgehog (SHH), a key signaling protein capable of regulating the differentiation process during retinal development, was assessed .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	WB analysis of MIO-M1 cells maintained under NG and HG conditions revealed a significant increase in SHH protein levels in the latter.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	IF observations confirmed such results and provided insights into the differential cytoplasmic localization of the protein between NG and HG MIO-M1 cells.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In the majority of NG MIO-M1 cells (about 85%), SHH was widespread throughout the cytoplasm.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In contrast, in approximately 70% of HG-MIO-M1 cells, although SHH maintained a widespread distribution, it was also predominantly localized in punctate structures (dot-like formations) within the cytoplasm and near the plasma membrane (Figure 6C,D).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	The WB results reported in Figure 7A,B show that SHH expression was significantly increased in NG MIO-M1 cells subjected to GF treatments (III–V), compared to their basal glucose treatment (I).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Although a clear trend towards increased SHH expression was also observed in response to sustained high-glucose treatment (II), this increase was not statically significant (Figure 7A,B).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Conversely, HG MIO-M1 cells subjected to treatments II and V showed no change in SHH expression compared to the basal glucose treatment, while a decrease of the SHH protein level in cells exposed to GF treatments III and IV was observed (Figure 7A,B).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	IF basically confirmed these findings at the protein level and revealed a distinct cytoplasm localization of the protein in NG MIO-M1 cells exposed to the different glucose treatments (II–V) compared to the basal glucose treatment (I).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In these cells, the increased level of SHH expression observed under sustained high-glucose (II) and GF (III–V) treatments was accompanied by a significant change in the intracellular localization of SHH, showing a more punctate pattern, consistent with that of HG MIO-M1 cells (Figure 7C,D).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In contrast, HG MIO-M1 cells exposed to all different glucose treatments showed no changes of intracellular SHH distribution compared to the basal glucose treatment (Figure 7C,D).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	These results underline the notion that Müller glia undergo adaptation or desensitization when cultured in a chronically hyperglycemic environment and subsequently subjected to further glucose-induced stress.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	IF observations and q-PCR analyses for SRY-Box Transcription Factor 2 (SOX2), a transcription factor that plays an important role in reprogramming Zebrafish Müller glia , showed expression changes of this transcription factor parallel to that of SHH.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In short, while approximately 55% of NG MIO-M1 cells were positive for the protein, in line with Lawrence et al. (2007) , this increased to 80% in HG MIO-M1 cells (Figure 8A,B).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	q-PCR analysis confirmed a higher SOX2 transcript in these latter in comparison to the former (Figure 8C).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Additionally, q-PCR analysis revealed a significant increase of SOX2 mRNA levels in NG MIO-M1 cells exposed to sustained high-glucose (II) and GF (III–V) treatments compared to the basal glucose treatment (I) (Figure 8D).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In contrast, no significant changes in SOX2 mRNA levels were observed in HG MIO-M1 cells when the cells were exposed to the same glucose treatments (Figure 8D).
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Müller glial cells play fundamental roles in retinal tissue functions.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Therefore, studies into their biology and functions may contribute to understanding the causes of retinal pathologies and to develop strategies to alleviate their outcomes.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	The aim of this study was to investigate the impact of high glucose and glucose fluctuations on critical cellular processes, such as gliosis and reprogramming in Müller cells.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	These processes are of pivotal importance in the onset and progression of retinal neurodegeneration and diabetic retinopathy.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In the present study, we observed a significant increase in GFAP expression in NG MIO-M1 cells exposed to sustained high-glucose and GF treatments, indicating a glucose stress-induced gliotic response.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	This response was associated with other markers of reactive gliosis, such as the morphological changes associated with the transition from a bipolar to a radial morphology and the overexpression/reorganization of vimentin intermediate filament.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	These findings indicate that NG MIO-M1 cells are sensitive to glucose changes in their surrounding environment, potentially functioning as a protective or reactive mechanism in response to metabolic stress.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	The remarkable sensitivity to glucose stress and the prompt reactive gliosis observed in MIO-M1 cells were found to be similar to those observed in Müller cells of diabetic subjects .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	This finding confirms the suitability of MIO-M1 cells as an in vitro model to study Müller cell response.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In contrast, HG MIO-M1 cells maintain a basal high level of both GFAP and Vimentin together to less organized intermediate filaments across the different glucose treatments, suggesting a limited and static response to sustained high-glucose and GF treatments.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	These findings highlight that MIO-M1 cells in chronic hyperglycemic conditions may adapt or develop enhanced tolerance over time due to constant and prolonged exposure to high glucose levels.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	The adaptation or desensitization of HG MIO-M1 cells to sustained high-glucose and GFs may indicate a saturation of the gliotic response, which is characteristic of prolonged hyperglycemic states, as observed in uncontrolled diabetes .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	This adaptation or desensitization may result in a reduction in the cellular capacity to respond to additional metabolic challenges or stress.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	To understand the clinical consequences of Müller cell gliosis it is essential to also consider the reprogramming process, which may include dedifferentiation of Müller cells and regeneration of various retinal neurons, contributing to retinal tissues repair under certain conditions.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In species such as birds , zebrafish , and rodents , Müller glia contribute as the primary source of retinal regeneration.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	When the retina is damaged, Müller glia can dedifferentiate into Müller glia-derived progenitor cells (MGPCs), acquiring a progenitor-like phenotype and starting to proliferate, thus contributing to retinal repair .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Notably, Müller glia in lower vertebrates exhibit a remarkable ability to regenerate retinal neurons, contrasting with the limited regenerative capacity seen in mammals, including humans , where Müller cells typically respond to injury with reactive gliosis, leading to scarring rather than regeneration .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	The SHH signaling pathway is essential for the proper development of all vertebrate retinas , and several studies have demonstrated the involvement of SHH signaling in the proliferation and differentiation of MGPCs, contributing to retinal regeneration in lower vertebrate .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	However, in mammals, the SHH pathway alone may not be sufficient to overcome the intrinsic limitations of Müller cells to regenerate neuronal cells.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In our study, we observed an increase in SHH expression in NG MIO-M1cells exposed to sustained high-glucose and GF treatments.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	This suggests that these cells are engaging in pathways associated with cellular dedifferentiation and potential reprogramming towards a progenitor state.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	The increased SHH protein expression in NG cells was accompanied by significant changes in the intracellular localization of SHH, exhibiting a more spotted/punctate pattern in the cytoplasm and in close proximity to the plasma membrane.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	These findings suggest an enhanced state of cellular activity with increased synthesis and potential accumulation of SHH, aligning with the findings in lower vertebrates and rodents, where increased SHH signaling plays a role in Müller cell reprogramming and retinal regeneration .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Furthermore, significantly higher levels of SHH were observed in HG MIO-M1 cells compared to NG MIO-M1 cells.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Conversely, HG MIO-M1 cells exposed to GF treatments showed a decrease in SHH protein expression levels.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	A trend toward a decrease was also observed in response to sustained high-glucose.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	This finding is consistent with a previous study in which the authors observed a decrease in SHH signaling in reactive astrocytes of the cerebral cortex after acute, focal injury, particularly in cells proximal to the lesion site .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	This highlights that the negative regulation of SHH activity in astrocytes is context-dependent and varies with the degree of cellular damage.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In our study, the decreased SHH levels observed in HG MIO-M1 cells exposed to GFs indicate that the severity of metabolic stress may influence SHH signaling pathways through a similar mechanism.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	These observations underscore the notion that glial cells, including Müller cells and astrocytes, exhibit differential SHH responses depending on the extent of glucose stress, highlighting the dynamic regulation of SHH signaling in glial cells.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Although the SHH level has decreased, the cells that express it maintain a dot-like distribution of SHH inside the cells.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Our findings suggest that, under NG condition, exposure to sustained high-glucose and GFs stimulates SHH expression, potentially promoting the dedifferentiation and reprogramming of Müller cells.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	However, in HG-adapted cells, additional metabolic stress from GF treatments leads to decreased SHH expression, possibly impairing regenerative capacity and enhancing gliotic responses.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	These observations indicate that the severity and fluctuation of glucose stress influence SHH signaling pathways, affecting the balance between neuroprotection and gliosis in the retina.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	A previous study demonstrated that high-glucose conditions have been associated with increased SOX2 levels in human Müller glial cells, which may support cell survival and regeneration under stress conditions .
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In our experiments, SOX2 was significantly upregulated in HG MIO-M1 cells, compared to NG MIO-M1 cells.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Furthermore, NG MIO-M1 cells have the capacity to modulate this gene when exposed to sustained high-glucose and GF treatments, exhibiting a consistent upregulation of SOX2.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	These findings are consistent with the observed upregulation of SHH in NG MIO-M1 cells exposed to different glucose treatments.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	Unlike NG MIO-M1cells, HG MIO-M1 cells exposed to the same glucose treatments do not show variations in SOX2 expression.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	This different behavior of HG MIO-M1 cells, with regards to SOX2 and SHH expression, may be due to the different roles and regulatory mechanisms in the glial cells of these two genes.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	SOX2 is a transcription factor crucial for maintaining stemness and promoting cell progenitor proliferation, and its upregulation could be a response to cellular stress in order to maintain or enhance regenerative capacity.
PMC11641291	Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.	In contrast, SHH signaling, which is involved in cell differentiation and tissue patterning, may be more sensitive to metabolic perturbations with a more dynamic regulation, which can lead to its downregulation under glucose fluctuations treatments.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

The hallmarks of diabetes-induced RN, which include neural-cell apoptosis, diminished retinal neuronal function, and reactive gliosis, have been observed to occur prior to the onset of overt microangiopathy in experimental models of DR, diabetic patients, and post-mortem human retinas .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Müller Glia cells (MGs) are the main retinal glial cells, extending throughout the entire thickness from the inner to the outer limiting membranes, providing an anatomical link between the retinal neurons and the retinal blood vessels and thus responsible for the homeostatic and metabolic support of retinal nerve cells .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

MGs also play a pivotal role in maintaining neuronal health through the recycling of neurotransmitters and the prevention of neurotoxicity resulting from glutamate excess.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Additionally, they regulate ionic balance by buffering K ions and supply lactate as an energy source for photoreceptors .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In response to diabetic stress, MGs undergo morphological changes and exhibit an increased expression of the intermediate filament proteins, such as glial fibrillary acidic protein (GFAP), a key marker of reactive gliosis .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Reactive gliosis plays a dual role in the context of retinal damage, with the potential to both protect against damage and contribute to its progression in a diabetic milieu.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

The formation of a glial scar, while stabilizing the tissue structure, may also impair neuronal function through the release of proinflammatory mediators .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

The ambivalent role of MGs gliosis in DR underscores the complexity of their involvement, thereby emphasizing the necessity for a comprehensive understanding of the regulatory mechanisms involved.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

The impact of fluctuating glycemic conditions on MGs responses introduces an additional layer of complexity.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Fluctuations in blood glucose levels have been demonstrated to trigger diverse signaling pathways within MGs, resulting in an increase of the inner nuclear layer (INL) in subjects with diabetes , potentially leading to differential activation states or pathological outcomes .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

As a matter of fact, in recent years significant advancements have been made in understanding the pathophysiological roles of MGs in DR .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Studies in lower vertebrates, including fish and amphibians, have demonstrated that, following reactive gliosis, the sonic hedgehog (SHH) signaling pathway is activated, thereby promoting the reprogramming of MGs into various retinal cell types and contributing to retinal regeneration .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

The capacity for retinal regeneration is notably limited in mammals, including humans.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In response to injury, MGs typically undergo a process of reactive gliosis, which often results in scarring rather than regeneration .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

To achieve significant regenerative processes, additional manipulations of the SHH pathway may be required .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Recent studies have identified the potential genes involved in reprogramming MGs into retinal neurons, such as Ascl1, Sox2, and Lin28, which can be modulated to stimulate functional neuronal regeneration .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Despite the advancement of knowledge regarding the neurogenic process, however, the existing literature lacks insights into the influence of glucose metabolism on these processes.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In particular, as far as we know, there is no evidence in the literature indicating the effect of glucose metabolism on retinal neurogenesis.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Our previous in vitro studies, involving rat retinal Müller cell line rMC-1, have demonstrated that these cells are activated in response to both high glucose levels and glucose fluctuations (GFs), and that this activation is associated with cellular increased levels of GFAP and aquaporin-4 (AQPs) .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

These findings contributed to elucidating the dynamic responses of Müller cells under varied glycaemic conditions, suggesting the existence of a complex regulatory mechanism sensitive to metabolic fluctuations.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

However, to better understand the mechanisms underlying DR in humans, it is essential to investigate whether or not similar responses occur in human Müller cell lines, as cellular responses may differ between animal models and humans, particularly due to differences in complexity and disease progression.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

The present study aims to fill this gap by investigating the gliotic response of human Müller cell line MIO-M1, a well-established model in human retinal research, exposed to sustained high-glucose and glucose fluctuation treatments that mimic the different glycemic conditions that can occur in diabetic patients.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Furthermore, the study aims to investigate the reprogramming potential of MIO-M1 cells under different glucose stress conditions, focusing on their potential for dedifferentiation and neurogenesis.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

These processes are of significant interest in understanding how glial cells respond to pathological conditions such as DR.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

This approach allowed one to explore not only the gliotic response but also the potential for reprogramming Müller cells into a more progenitor-like state, a key aspect for the development of future regenerative therapies.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

The results obtained might contribute to elucidating the dynamic responses of MGs under different glycemic conditions, suggesting the existence of a complex regulatory mechanism sensitive to glucose metabolic stress.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

WB analysis carried out on MIO-M1 cells cultured under HG condition showed a significant increase in GFAP levels compared to MIO-M1 cells cultured under NG condition (fold change of about 2 in HG vs. NG), indicating an enhanced gliotic response under hyperglycemic stress conditions (Figure 1A,B).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Such significant increases of GFAP levels was also observed when NG MIO-M1 cells were exposed to sustained high-glucose (II) and GF (III) treatments compared to their basal glucose (I) (Figure 2A,B and Figure S1).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Although a clear trend towards increased GFAP expression was also observed across GF treatments IV and V, these increases were not statistically significant (Figure 2A,B).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In contrast, MIO-M1 cells cultured under HG condition showed a markedly different response, with no significant changes in GFAP levels when these cells were exposed to sustained high-glucose (II) and GF (III–V) treatments compared to their basal glucose treatment (I) (Figure 2A,B).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

The impact of the experimental culture conditions described above on GFAP expression in MIO-MI cells was also investigated by IF, and cellular bipolar or radial morphologies, typical of quiescent and activated Müller cells, respectively , were also recorded.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Under NG condition, GFAP expression was observed in a limited number of MIO-MI cells, approximately 15%, which were GFAP positive, consistent with literature data .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Typical of low active Müller cell status, GFAP staining in NG MIO-M1 cells was faint, while bipolar or radial morphologies were equally distributed (Figure 3A,B).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In contrast, MIO-M1 cells maintained in the HG condition showed intense GFAP staining, with almost 80% displaying radial morphology (Figure 3A,B), indicating that these cells are in a more activated state.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Similarly to the WB results, in NG MIO-M1cells, subjected to all different glucose treatments (II–V), a significant increase in the GFAP intensity and radial morphology were observed, compared to the basal glucose treatment (I) (NG: 47.85% condition I vs. 69,63% condition II; 77.80% condition III; 74.21% condition IV; 81.42% condition V) (Figure 4A,B).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In contrast, in HG MIO-M1 cells subjected to any experimental glucose treatments, no significant change in GFAP positivity or intensity and morphology were detected compared to the basal glucose treatment (Figure 4A,C).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

IF for vimentin filaments, another well-established marker of Müller cell stress response, on MIO-M1 cells subjected to all different glucose treatments gave similar results to those described for GFAP and cell morphology changes.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Briefly, NG MIO-M1 cells in the basal glucose treatment (I) exhibited intact cytoskeletal integrity, with well-organized and linear vimentin filaments.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Following the exposure of NG MIO-M1 cells to sustained high-glucose (II) and GF (III–V) treatments, a significant increase in vimentin expression was observed, compared to the basal glucose treatment (I).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

This increase was associated with a more disorganized appearance of vimentin filaments, reflecting increased cellular stress and cytoskeletal reorganization (Figure 5).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Such a result was also observed in HG MIO-M1 cells maintained in the basal glucose treatment (I) and when these cells were exposed to all different glucose treatments (II–V) (Figure 5).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In order to investigate the activation of the neurogenic potential of MIO-M1 cells under the various glucose treatments investigated here, the expression and cellular localization of sonic hedgehog (SHH), a key signaling protein capable of regulating the differentiation process during retinal development, was assessed .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

WB analysis of MIO-M1 cells maintained under NG and HG conditions revealed a significant increase in SHH protein levels in the latter.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

IF observations confirmed such results and provided insights into the differential cytoplasmic localization of the protein between NG and HG MIO-M1 cells.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In the majority of NG MIO-M1 cells (about 85%), SHH was widespread throughout the cytoplasm.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In contrast, in approximately 70% of HG-MIO-M1 cells, although SHH maintained a widespread distribution, it was also predominantly localized in punctate structures (dot-like formations) within the cytoplasm and near the plasma membrane (Figure 6C,D).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

The WB results reported in Figure 7A,B show that SHH expression was significantly increased in NG MIO-M1 cells subjected to GF treatments (III–V), compared to their basal glucose treatment (I).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Although a clear trend towards increased SHH expression was also observed in response to sustained high-glucose treatment (II), this increase was not statically significant (Figure 7A,B).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Conversely, HG MIO-M1 cells subjected to treatments II and V showed no change in SHH expression compared to the basal glucose treatment, while a decrease of the SHH protein level in cells exposed to GF treatments III and IV was observed (Figure 7A,B).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

IF basically confirmed these findings at the protein level and revealed a distinct cytoplasm localization of the protein in NG MIO-M1 cells exposed to the different glucose treatments (II–V) compared to the basal glucose treatment (I).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In these cells, the increased level of SHH expression observed under sustained high-glucose (II) and GF (III–V) treatments was accompanied by a significant change in the intracellular localization of SHH, showing a more punctate pattern, consistent with that of HG MIO-M1 cells (Figure 7C,D).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In contrast, HG MIO-M1 cells exposed to all different glucose treatments showed no changes of intracellular SHH distribution compared to the basal glucose treatment (Figure 7C,D).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

These results underline the notion that Müller glia undergo adaptation or desensitization when cultured in a chronically hyperglycemic environment and subsequently subjected to further glucose-induced stress.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

IF observations and q-PCR analyses for SRY-Box Transcription Factor 2 (SOX2), a transcription factor that plays an important role in reprogramming Zebrafish Müller glia , showed expression changes of this transcription factor parallel to that of SHH.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In short, while approximately 55% of NG MIO-M1 cells were positive for the protein, in line with Lawrence et al. (2007) , this increased to 80% in HG MIO-M1 cells (Figure 8A,B).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

q-PCR analysis confirmed a higher SOX2 transcript in these latter in comparison to the former (Figure 8C).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Additionally, q-PCR analysis revealed a significant increase of SOX2 mRNA levels in NG MIO-M1 cells exposed to sustained high-glucose (II) and GF (III–V) treatments compared to the basal glucose treatment (I) (Figure 8D).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In contrast, no significant changes in SOX2 mRNA levels were observed in HG MIO-M1 cells when the cells were exposed to the same glucose treatments (Figure 8D).

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Müller glial cells play fundamental roles in retinal tissue functions.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Therefore, studies into their biology and functions may contribute to understanding the causes of retinal pathologies and to develop strategies to alleviate their outcomes.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

The aim of this study was to investigate the impact of high glucose and glucose fluctuations on critical cellular processes, such as gliosis and reprogramming in Müller cells.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

These processes are of pivotal importance in the onset and progression of retinal neurodegeneration and diabetic retinopathy.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In the present study, we observed a significant increase in GFAP expression in NG MIO-M1 cells exposed to sustained high-glucose and GF treatments, indicating a glucose stress-induced gliotic response.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

This response was associated with other markers of reactive gliosis, such as the morphological changes associated with the transition from a bipolar to a radial morphology and the overexpression/reorganization of vimentin intermediate filament.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

These findings indicate that NG MIO-M1 cells are sensitive to glucose changes in their surrounding environment, potentially functioning as a protective or reactive mechanism in response to metabolic stress.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

The remarkable sensitivity to glucose stress and the prompt reactive gliosis observed in MIO-M1 cells were found to be similar to those observed in Müller cells of diabetic subjects .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

This finding confirms the suitability of MIO-M1 cells as an in vitro model to study Müller cell response.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In contrast, HG MIO-M1 cells maintain a basal high level of both GFAP and Vimentin together to less organized intermediate filaments across the different glucose treatments, suggesting a limited and static response to sustained high-glucose and GF treatments.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

These findings highlight that MIO-M1 cells in chronic hyperglycemic conditions may adapt or develop enhanced tolerance over time due to constant and prolonged exposure to high glucose levels.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

The adaptation or desensitization of HG MIO-M1 cells to sustained high-glucose and GFs may indicate a saturation of the gliotic response, which is characteristic of prolonged hyperglycemic states, as observed in uncontrolled diabetes .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

This adaptation or desensitization may result in a reduction in the cellular capacity to respond to additional metabolic challenges or stress.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

To understand the clinical consequences of Müller cell gliosis it is essential to also consider the reprogramming process, which may include dedifferentiation of Müller cells and regeneration of various retinal neurons, contributing to retinal tissues repair under certain conditions.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In species such as birds , zebrafish , and rodents , Müller glia contribute as the primary source of retinal regeneration.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

When the retina is damaged, Müller glia can dedifferentiate into Müller glia-derived progenitor cells (MGPCs), acquiring a progenitor-like phenotype and starting to proliferate, thus contributing to retinal repair .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Notably, Müller glia in lower vertebrates exhibit a remarkable ability to regenerate retinal neurons, contrasting with the limited regenerative capacity seen in mammals, including humans , where Müller cells typically respond to injury with reactive gliosis, leading to scarring rather than regeneration .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

The SHH signaling pathway is essential for the proper development of all vertebrate retinas , and several studies have demonstrated the involvement of SHH signaling in the proliferation and differentiation of MGPCs, contributing to retinal regeneration in lower vertebrate .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

However, in mammals, the SHH pathway alone may not be sufficient to overcome the intrinsic limitations of Müller cells to regenerate neuronal cells.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In our study, we observed an increase in SHH expression in NG MIO-M1cells exposed to sustained high-glucose and GF treatments.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

This suggests that these cells are engaging in pathways associated with cellular dedifferentiation and potential reprogramming towards a progenitor state.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

The increased SHH protein expression in NG cells was accompanied by significant changes in the intracellular localization of SHH, exhibiting a more spotted/punctate pattern in the cytoplasm and in close proximity to the plasma membrane.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

These findings suggest an enhanced state of cellular activity with increased synthesis and potential accumulation of SHH, aligning with the findings in lower vertebrates and rodents, where increased SHH signaling plays a role in Müller cell reprogramming and retinal regeneration .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Furthermore, significantly higher levels of SHH were observed in HG MIO-M1 cells compared to NG MIO-M1 cells.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Conversely, HG MIO-M1 cells exposed to GF treatments showed a decrease in SHH protein expression levels.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

A trend toward a decrease was also observed in response to sustained high-glucose.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

This finding is consistent with a previous study in which the authors observed a decrease in SHH signaling in reactive astrocytes of the cerebral cortex after acute, focal injury, particularly in cells proximal to the lesion site .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

This highlights that the negative regulation of SHH activity in astrocytes is context-dependent and varies with the degree of cellular damage.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In our study, the decreased SHH levels observed in HG MIO-M1 cells exposed to GFs indicate that the severity of metabolic stress may influence SHH signaling pathways through a similar mechanism.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

These observations underscore the notion that glial cells, including Müller cells and astrocytes, exhibit differential SHH responses depending on the extent of glucose stress, highlighting the dynamic regulation of SHH signaling in glial cells.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Although the SHH level has decreased, the cells that express it maintain a dot-like distribution of SHH inside the cells.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Our findings suggest that, under NG condition, exposure to sustained high-glucose and GFs stimulates SHH expression, potentially promoting the dedifferentiation and reprogramming of Müller cells.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

However, in HG-adapted cells, additional metabolic stress from GF treatments leads to decreased SHH expression, possibly impairing regenerative capacity and enhancing gliotic responses.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

These observations indicate that the severity and fluctuation of glucose stress influence SHH signaling pathways, affecting the balance between neuroprotection and gliosis in the retina.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

A previous study demonstrated that high-glucose conditions have been associated with increased SOX2 levels in human Müller glial cells, which may support cell survival and regeneration under stress conditions .

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In our experiments, SOX2 was significantly upregulated in HG MIO-M1 cells, compared to NG MIO-M1 cells.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Furthermore, NG MIO-M1 cells have the capacity to modulate this gene when exposed to sustained high-glucose and GF treatments, exhibiting a consistent upregulation of SOX2.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

These findings are consistent with the observed upregulation of SHH in NG MIO-M1 cells exposed to different glucose treatments.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

Unlike NG MIO-M1cells, HG MIO-M1 cells exposed to the same glucose treatments do not show variations in SOX2 expression.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

This different behavior of HG MIO-M1 cells, with regards to SOX2 and SHH expression, may be due to the different roles and regulatory mechanisms in the glial cells of these two genes.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

SOX2 is a transcription factor crucial for maintaining stemness and promoting cell progenitor proliferation, and its upregulation could be a response to cellular stress in order to maintain or enhance regenerative capacity.

PMC11641291

Gliotic Response and Reprogramming Potential of Human Müller Cell Line MIO-M1 Exposed to High Glucose and Glucose Fluctuations.

In contrast, SHH signaling, which is involved in cell differentiation and tissue patterning, may be more sensitive to metabolic perturbations with a more dynamic regulation, which can lead to its downregulation under glucose fluctuations treatments.