Noncanonical CDK4 signaling rescues diabetes in a mouse model by promoting ß cell differentiation.

Expanding ß cell mass is a critical goal in the fight against diabetes. CDK4, an extensively characterized cell cycle activator, is required to establish and maintain ß cell number. ß cell failure in the IRS2-deletion mouse type 2 diabetes model is, in part, due to loss of CDK4 regulator cyclin D2. We set out to determine whether replacement of endogenous CDK4 with the inhibitor-resistant mutant CDK4-R24C rescued the loss of ß cell mass in IRS2-deficient mice. Surprisingly, not only ß cell mass but also ß cell dedifferentiation was effectively rescued, despite no improvement in whole body insulin sensitivity. Ex vivo studies in primary islet cells revealed a mechanism in which CDK4 intervened downstream in the insulin signaling pathway to prevent FOXO1-mediated transcriptional repression of critical ß cell transcription factor Pdx1. FOXO1 inhibition was not related to E2F1 activity, to FOXO1 phosphorylation, or even to FOXO1 subcellular localization, but rather was related to deacetylation and reduced FOXO1 abundance. Taken together, these results demonstrate a differentiation-promoting activity of the classical cell cycle activator CDK4 and support the concept that ß cell mass can be expanded without compromising function.

Endoplasmic Reticulum Stress Induced Proliferation Remains Intact in Aging Mouse ß-Cells.

Aging is associated with loss of proliferation of the insulin-secreting ß-cell, a possible contributing factor to the increased prevalence of type 2 diabetes in the elderly. Our group previously discovered that moderate endoplasmic reticulum (ER) stress occurring during glucose exposure increases the adaptive ß-cell proliferation response. Specifically, the ATF6α arm of the tripartite Unfolded Protein Response (UPR) promotes ß-cell replication in glucose excess conditions. We hypothesized that ß-cells from older mice have reduced proliferation due to aberrant UPR signaling or an impaired proliferative response to ER stress or ATF6α activation. To investigate, young and old mouse islet cells were exposed to high glucose with low-dose thapsigargin or activation of overexpressed ATF6α, and ß-cell proliferation was quantified by BrdU incorporation. UPR pathway activation was compared by qPCR of target genes and semi-quantitative Xbp1 splicing assay. Intriguingly, although old ß-cells had reduced proliferation in high glucose compared to young ß-cells, UPR activation and induction of proliferation in response to low-dose thapsigargin or ATF6α activation in high glucose were largely similar between young and old. These results suggest that loss of UPR-led adaptive proliferation does not explain the reduced cell cycle entry in old ß-cells, and raise the exciting possibility that future therapies that engage adaptive UPR could increase ß-cell number through proliferation even in older individuals.

Intersection of the ATF6 and XBP1 ER stress pathways in mouse islet cells.

Success or failure of pancreatic beta cell adaptation to ER stress is a determinant of diabetes susceptibility. The ATF6 and IRE1/XBP1 pathways are separate ER stress-response effectors important to beta cell health and function. ATF6α. and XBP1 direct overlapping transcriptional responses in some cell types. However, the signaling dynamics and interdependence of ATF6α and XBP1 in pancreatic beta cells have not been explored. To assess pathway-specific signal onset, we performed timed exposures of primary mouse islet cells to ER stressors and measured the early transcriptional response. Comparing the time course of induction of ATF6 and XBP1 targets suggested that the two pathways have similar response dynamics. The role of ATF6α in target induction was assessed by acute knockdown using islet cells from Atf6α flox/flox mice transduced with adenovirus expressing Cre recombinase. Surprisingly, given the mild impact of chronic deletion in mice, acute ATF6α knockdown markedly reduced ATF6-pathway target gene expression under both basal and stressed conditions. Intriguingly, although ATF6α knockdown did not alter Xbp1 splicing dynamics or intensity, it did reduce induction of XBP1 targets. Inhibition of Xbp1 splicing did not decrease induction of ATF6α targets. Taken together, these data suggest that the XBP1 and ATF6 pathways are simultaneously activated in islet cells in response to acute stress and that ATF6α is required for full activation of XBP1 targets, but XBP1 is not required for activation of ATF6α targets. These observations improve understanding of the ER stress transcriptional response in pancreatic islets.

Prevalence and pattern of amblyopia in a rural hospital in Ghana.

Amblyopia is a developmental ocular disease of childhood-onset which may lead to persistent sequelae into adulthood. Early detection and management of amblyopia usually result in an improved visual outcome. The purpose of this study was to determine the prevalence and pattern of amblyopia in a rural hospital in Ghana. Clinical records of patients seen (from January 2014 to December 2018) at Westphalian Medical Center, Oyoko, Ashanti Region, Ghana, were reviewed retrospectively. Unilateral amblyopia was defined as a two-line interocular difference or more in visual acuity. Bilateral amblyopia was defined as best-corrected visual acuity (BCVA) of Snellen 6/12 or worse in both eyes, with evidence of bilateral ametropia or obstruction of the visual pathway. Following a review of 12,602 patient records, 258 cases of amblyopia were identified. The mean (±SD) presenting age of amblyopic patients was 24.3 ± 16.1 years, with a male-to-female ratio of 1:1.1. The period prevalence of amblyopia was 2.04%. The period prevalence of unilateral and bilateral amblyopia was 1.38% and 0.66%, respectively. The most prevalent form of amblyopia was refractive with a cumulative prevalence of 1.42%. Strabismic and stimulus deprivation amblyopia accounted for 0.36% and 0.21% of all amblyopic cases, respectively. A major cause of amblyopia in this population was refractive error, hence the use of spectacle correction for its initial management. Repeated assessment after an appropriate period of refractive adaptation would elucidate the proportion of amblyopias needing additional treatment modalities. Vision screening for early detection of amblyopia in childhood with accessible and effective management of amblyopia (including refractive correction and occlusion treatment) is necessary to reduce the impact of amblyopia in Ghana.

Assessment of availability, awareness and perception of stakeholders regarding preschool vision screening in Kumasi, Ghana: An exploratory study.

BACKGROUND: Regardless of the importance of preschool vision screening (PSVS), there is limited data on the current state of these programs in Africa (particularly Ghana). This study sought to investigate the level of awareness and perception of stakeholders regarding PSVS, its availability and related policies/programmes in the Kumasi Metropolis, Ghana. METHODS: This descriptive cross-sectional study included 100 systematically sampled preschools in the metropolis (using probability proportional-to-size method); 72 private schools and 28 public schools. Convenience sampling was used to recruit stakeholders of preschools (teachers, head teachers, proprietors, administrators, directors, and educationists), and were interviewed using a well-structured questionnaire. Questionnaires were administered to all eligible respondents who were present at the time of data collection. RESULTS: A total of 344 respondents participated in the study; 123 (35.8%) males and 221 (64.2%) females. The overall mean age of respondents was 37.63 ±12.20 years (18-71 years). Of the respondents, 215 (62.5%), 94 (27.3%), and 35 (10.2%) were enrolled from private schools, public schools, and Metropolitan Education Directorate, respectively. 73.8% of respondents reported the absence of routine PSVS in schools whereas 90.1% reported no written policies for PSVS in schools. Only 63.6% of respondents were aware of PSVS whereas more than half (59.6%) of all respondents perceived PSVS to be very important for preschoolers. Private school ownership was significantly associated with availability of PSVS whereas age, teachers, private school ownership, and preschool experience > 10 years were significantly associated with awareness of PSVS (P < 0.05). However, there was no significant association between sociodemographic factors and perception of PSVS. CONCLUSION: PSVS is largely unavailable in most Ghanaian schools. Majority of stakeholders were aware of PSVS and agreed to its implementation and incorporation into schools' health programmes. There is the need to implement a national programme/policy on preschool vision screening in Ghana.

DNA Damage Does Not Cause BrdU Labeling of Mouse or Human ß-Cells.

Pancreatic ß-cell regeneration, the therapeutic expansion of ß-cell number to reverse diabetes, is an important goal. Replication of differentiated insulin-producing cells is the major source of new ß-cells in adult mice and juvenile humans. Nucleoside analogs such as BrdU, which are incorporated into DNA during S-phase, have been widely used to quantify ß-cell proliferation. However, reports of ß-cell nuclei labeling with both BrdU and γ-phosphorylated H2A histone family member X (γH2AX), a DNA damage marker, have raised questions about the fidelity of BrdU to label S-phase, especially during conditions when DNA damage is present. We performed experiments to clarify the causes of BrdU-γH2AX double labeling in mouse and human ß-cells. BrdU-γH2AX colabeling is neither an age-related phenomenon nor limited to human ß-cells. DNA damage suppressed BrdU labeling and BrdU-γH2AX colabeling. In dispersed islet cells, but not in intact islets or in vivo, pro-proliferative conditions promoted both BrdU and γH2AX labeling, which could indicate DNA damage, DNA replication stress, or cell cycle-related intrinsic H2AX phosphorylation. Strategies to increase ß-cell number must not only tackle the difficult challenge of enticing a quiescent cell to enter the cell cycle, but also achieve safe completion of the cell division process.