Pax4 Research Articles

Rosa canina extract relieves methylation alterations of pancreatic genes in STZ-induced diabetic rats : Gene methylation in diabetic rats treated with an extract.

Diabetes is a chronic metabolic disease that affects many parts of the body. Considering diabetes as a beta cells' defect and loss, the focus is on finding mechanisms and compounds involved in stimulating the function and regeneration of pancreatic β-cells. DNA methylation as an epigenetic mechanism plays a pivotal role in the β-cells' function and development. Considering the regenerative and anti-diabetic effects of Rosa canina extract, this study aimed to assess the methylation levels of Pdx-1, Pax-4, and Ins-1 genes in diabetic rats treated with Rosa Canina extract. Streptozotocin-induced diabetic rats were used to evaluate the frequency of Pdx-1, Pax-4, and Ins-1 gene methylation. Treatment groups were exposed to Rosa canina as spray-dried and decoction extracts. Following blood glucose measurement, pancreatic DNA was extracted and bisulfited. Genes' methylation was measured using MSP-PCR and qRT-PCR techniques. Oral administration of Rosa canina extracts significantly reduced blood sugar levels in diabetic rats compared to the control group. The methylation levels of the Pdx-1, Pax-4, and Ins-1 genes promoter in streptozotocin-induced diabetic rats increased compared to the control rats while, the treatment of diabetic rats with Rosa canina extracts, spray-dried samples especially, led to a decreased methylation in these genes. The results of this study showed that Rosa canina extract as a spray-dried sample could be effective in treating diabetes by regulating the methylation of genes including Pdx-1, Pax-4, and Ins-1 involved in the activity and regeneration of pancreatic islet cells.

PAX4 gene delivery improves β-cell function in human islets of Type II diabetes.

Aim: Type II diabetes (T2D) stems from insulin resistance, with β-cell dysfunction as a hallmark in its progression. Studies reveal that β cells undergo apoptosis or dedifferentiation during T2D development. The transcription factor PAX4 is vital for β differentiation and survival, thus may be a potential enhancer of β-cell function in T2D islets. Materials & methods: Human PAX4 cDNA was delivered into T2D human islets with an adenoviral vector, and its effects on β cells were examined. Results: PAX4 gene delivery significantly improved β-cell survival, and increased β-cell composition in the T2D human islets. Basal insulin and glucose-stimulated insulin secretion in PAX4-expressing islets were substantially higher than untreated or control-treated T2D human islets. Conclusion: Introduced PAX4 expression in T2D human islets improves β-cell function, thus could provide therapeutic benefits for T2D treatment.

In Silico Functional and Structural Analysis of Non-synonymous Single Nucleotide Polymorphisms (nsSNPs) in Human Paired Box 4 Gene.

In human genome, members of Paired box (PAX) transcription factor family are highly sequence-specific DNA-binding proteins. Among PAX gene family members, PAX4 gene has significant role in growth, proliferation, differentiation, and insulin secretion of pancreatic β-cells. Single nucleotide polymorphisms (SNPs) in PAX4 gene progress in the pathogenesis of various human diseases. Hence, the molecular mechanism of how these SNPs in PAX4 gene significantly progress diseases pathogenesis needs to be elucidated. For the reason, a series of bioinformatic analyzes were done to identify the SNPs of PAX4 gene that contribute in diseases pathogenesis. From the analyzes, 4145 SNPs (rsIDs) in PAX4 gene were obtained, where, 362 missense (8.73%), 169 synonymous (4.08%), and 2323 intron variants (56.04%). The rest SNPs were unspecified. Among the 362 missense variants, 118 nsSNPs were found as deleterious in SIFT analysis. Among those, 25 nsSNPs were most probably damaging and 23 were deleterious as observed in PolyPhen-2 and PROVEAN analyzes, respectively. Following all analyzes, 14 nsSNPs (rs149708455, rs115887120, rs147279315, rs35155575, rs370095957, rs373939873, rs145468905, rs121917718, rs2233580, rs3824004, rs372751660, rs369459316, rs375472849, rs372497946) were common and observed as deleterious, probably damaging, affective and diseases associated. Following structural analyzes, 11 nsSNPs guided proteins were found as most unstable and highly conserved. Among these, R20W, R39Q, R45Q, R60H, G65D, and A223D mutated proteins were highly harmful. Hence, the results from above-mentioned integrated comprehensive bioinformatic analyzes guide how different nsSNPs in PAX4 gene alter structural and functional characteristics of the protein that might progress diseases pathogenesis in human including type 2 diabetes.

A Novel Variant in the PAX4 Gene Causes Maturity-Onset Diabetes of the Young (MODY), Type IX with Neurodevelopmental Disorder

A genetically diverse condition, maturity-onset diabetes of the young (MODY), frequently develops before the age of 25. MODY is caused by disease-causing sequence variations in the PAX4 gene, which is found on chromosome 7q32.1. Additionally, it has also been observed that variants in PAX4 have also been associated with neurodevelopmental disability. Whole exome sequencing (WES) followed by Sanger sequencing was performed for all the available affected and unaffected members of the family. Data analysis revealed a novel heterozygous nonsense variant (c.61C>T; p.Gln21*) in the PAX4 gene in the affected individuals, which segregated perfectly with the disease phenotype. The present study adds to the PAX4 mutation spectrum and reports on the first case of MODY associated with neurodevelopmental disorders in humans.

Метод секвенирования нового поколения в дифференциальной диагностике сахарного диабета у детей

In addition to type 1 and 2 diabetes mellitus (DM), endocrine disorders in children include hereditary forms of diabetes, such as maturity onset diabetes of the young (MODY). Since pathogenic mechanisms of chronic hyperglycemia in MODY are different from those in DM1 and DM2, it requires other therapeutic approaches. The diagnosis of MODY is confirmed by expensive molecular testing, such as next generation sequencing (NGS). Therefore, the strategy of choosing candidates for NGS is very important. Objective. To optimize the algorithm of choosing patients for NGS testing for DM type verification. Patients and methods. This study included 97 patients aged 1–18 years suspected of having MODY. We used NGS to confirm the diagnosis and identify polymorphisms in MODY-associated genes. Results. Fifty-three patients were found to have polymorphisms in MODY-associated genes, including GCK gene (MODY2) (n = 44), NHF1A gene (MODY3) (n = 8), and PAX4 gene (MODY9) (n = 1). Comparison of family histories of patients with MODY-associated polymorphisms and those in whom clinical diagnosis of MODY was not confirmed by NGS demonstrated significant differences: children with verified MODY were more likely to have first-degree relatives with some carbohydrate metabolism disorder (CMDs). Clinical, laboratory, and anthropometric parameters, as well as levels of insulin secretion and carbohydrate metabolism were similar in both groups. However, patients with non-confirmed MODY received insulin therapy more frequently than those with verified MODY as the majority of them were on a diet. Conclusion. NGS confirms the diagnosis of MODY in patients with different CMDs. A tailored approach should be used to choose patients for NGS to confirm MODY. Key words: monogenic diabetes mellitus in children, diabetes mellitus, next generation sequencing, GCK, HNF1A, MODY

Hepatocyte Nuclear Factor 4-α (HNF4α) controls the insulin resistance-induced pancreatic β-cell mass expansion

BackgroundRegardless of the etiology, any type of DM presents a reduction of insulin-secreting cell mass, so it is important to investigate pathways that induce the increase of this cell mass. AimBased on the fact that (1) HNF4α is crucial for β-cell proliferation, (2) DEX-induced IR promotes β-cell mass expansion, and (3) the stimulation of β-cell mass expansion may be an important target for DM therapies, we aimed to investigate whether DEX-induced proliferation of β pancreatic cells is dependent on HNF4α. MethodsWe used WildType (WT) and Knockout (KO) mice for HNF4-α, treated or not with 100 mg/Kg/day of DEX, for 5 consecutive days. One day after the last injection of DEX the IR was confirmed by ipITT and the mice were euthanized for pancreas removal. ResultsIn comparison to WT, KO mice presented increased glucose tolerance, lower fasting glucose and increased glucose-stimulates insulin secretion (GSIS). DEX induced IR in both KO and WT mice. In addition, DEX-induced β-cell mass expansion and an increase in the Ki67 immunostaining were observed only in WT mice, evidencing that IR-induced β-cell mass expansion is dependent on HNF4α. Also, we observed that DEX-treatment, in an HNF4α-dependent way, promoted an increase in PDX1, PAX4 and NGN3 gene expression. ConclusionsOur results strongly suggest that DEX-induced IR promotes β-cell mass expansion through processes of proliferation and neogenesis that depend on the HNF4α activity, pointing to HNF4α as a possible therapeutic target in DM treatment.

Identification of the First PAX4-MODY Family Reported in Brazil.

PurposeThe aim of this study was to sequence the coding region of the PAX4 gene in a Brazilian cohort with clinical manifestations of monogenic diabetes.Patients and MethodsThis study included 31 patients with autosomal dominant history of diabetes, age at diagnosis ≤40 years, BMI <30 kg/m2, and no mutations in GCK or HNF1A, HNF4A, and HNF1B. Screening of the PAX4 coding region was performed by Sanger sequencing. In silico algorithms were used to assess the potential impact of amino acid substitutions on protein structure and function. Additionally, PAX4-MODY family members and 158 control subjects without diabetes were analyzed for the identified mutation.ResultsThe molecular analysis of PAX4 has detected one missense mutation, p.Arg164Gln (c.491G>A), segregating with diabetes in a large Brazilian family. The mutation was absent among the control group. The index case is a woman diagnosed at 32 years of age with polyneuropathy and treated with insulin. She did not present diabetic renal disease or retinopathy. Family members with the PAX4 p.Arg164Gln mutation have a heterogeneous clinical manifestation and treatment response, with age at diagnosis ranging from 24 years to 50 years.ConclusionTo the best of our knowledge, this is the first study to report a PAX4-MODY family in Brazil. The age of PAX4-MODY diagnosis in the Brazilian family seems to be higher than the classical criteria for MODY. Our results reinforce the importance of screening large monogenic diabetes families for the understanding of the clinical manifestations of rare forms of diabetes for the specific and personalized treatment.

Pax4 Gene Delivery Improves Islet Transplantation Efficacy by Promoting β Cell Survival and α-to-β Cell Transdifferentiation

The transcription factor Pax4 plays an essential role in the development of insulin-producing β cells in pancreatic islets. Ectopic Pax4 expression not only promotes β cell survival but also induces α-to-β cell transdifferentiation. This dual functionality of Pax4 makes it an appealing therapeutic target for the treatment of insulin-deficient type 1 diabetes (T1D). In this study, we demonstrated that Pax4 gene delivery by an adenoviral vector, Ad5.Pax4, improved β cell function of mouse and human islets by promoting islet cell survival and α-to-β cell transdifferentiation, as assessed by multiple viability assays and lineage-tracing analysis. We then explored the therapeutic benefits of Pax4 gene delivery in the context of islet transplantation using T1D mouse models. Both mouse-to-mouse and human-to-mouse islet transplantation, via either kidney capsule or portal vein, were examined. In all settings, Ad5.Pax4-treated donor islets (mouse or human) showed substantially better therapeutic outcomes. These results suggest that Pax4 gene delivery into donor islets may be considered as an adjunct therapy for islet transplantation, which can either improve the therapeutic outcome of islet transplantation using the same amount of donor islets or allow the use of fewer donor islets to achieve normoglycemia.

Alternative Variants of Pax4 Human Transcription Factor: Comparative Transcriptional Activity

MODY is a group of genetically and clinically heterogeneous forms of diabetes characterized by auto-somal dominant inheritance and is subdivided in 13 subtypes dependent on the gene involved. The subtype MODY9 is a very rare form caused by mutations in the gene encoding the PAX4 transcription factor which is engaged in differentiation of pancreatic beta-cells. PAX4 contains two DNA-binding domains-Paired and Homeo. Expression of the human PAX4 gene is tissue-specific. The alternatively spliced mRNA variants encode for protein isoforms which differ within their N- and C-terminal regions. In this study, the transcriptional activities of the human PAX4 variants, both known and new ones, were determined. The full-length PAX4 containing intact DNA-binding domains was found to have maximal activity in transient expression system of the firefly luciferase reporter gene under control of the insulin promoter in HEK293 cells. The transcriptional activity is significantly reduced in the variants lacking eight N-terminal amino acid residues and/or variants whose Homeo domain is truncated from the C-terminus. Similar data were obtained with the glucagon promoter reporter system. The aberrant PAX4 variants were shown to retain stability and nuclear localization.

Phenolic-enriched blueberry-leaf extract attenuates glucose homeostasis, pancreatic β-cell function, and insulin sensitivity in high-fat diet–induced diabetic mice

Blueberry fruit exhibits strong antioxidant activity owing to the presence of anthocyanin. As blueberry-leaf extract (BLE) contains chlorogenic acid and flavonol glycosides, we hypothesized that phenolic-enriched BLE would improve glucose homeostasis and insulin sensitivity. In this study, we examined whether BLE administration decreases the glucose levels and enhances the pancreatic function in mice with high-fat diet (HFD)-induced obesity and diabetes. C57BL/6J mice were divided into the following four groups: control diet (CD), HFD (60 kcal% fat diet), BLE (HFD with 1% BLE wt/wt diet), and yerba mate extract (YME; HFD with 0.5% YME wt/wt diet). Dietary BLE and YME reduced glucose tolerance, body weight, and plasma glucose, glycated hemoglobin, insulin, homeostasis model assessment of insulin resistance, triglyceride (TG), and non-esterified fatty acid levels. Compared with those of the HFD group, BLE was found to significantly reduce the pancreatic islet size and insulin content. Moreover, it increased the mRNA levels of pancreatic β-cell proliferation-related genes, Ngn3, MafA, Pax4, Ins1, and Ins2, and pancreatic insulin signaling-related genes, IRS-1, IRS-2, PIK3ca, PDK1, PKCε, and GLUT-2, and decreased the transcriptional expression of the β-cell apoptosis-related gene, FoxO1. Both BLE and YME improved insulin sensitivity by inhibiting TG synthesis and enhancing lipid utilization in the liver and white adipose tissue (WAT). In pancreatic MIN6 β-cells, BLE and its main component (chlorogenic acid) increased β-cell proliferation and promoted insulin signaling. Overall, BLE enriched with phenolic compounds has the capacity to prevent HFD-induced glucose tolerance and hyperglycemia by improving the pancreatic β-cell function.

Insulin acts as a repressive factor to inhibit the ability of PAR2 to induce islet cell transdifferentiation

ABSTRACTRecently, we showed that pancreatitis in the context of profound β-cell deficiency was sufficient to induce islet cell transdifferentiation. In some circumstances, this effect was sufficient to result in recovery from severe diabetes. More recently, we showed that the molecular mechanism by which pancreatitis induced β-cell neogenesis by transdifferentiation was activation of an atypical GPCR called Protease-Activated Receptor 2 (PAR2). However, the ability of PAR2 to induce transdifferentiation occurred only in the setting of profound β-cell deficiency, implying the existence of a repressive factor from those cells. Here we show that the repressor from β-cells is insulin. Treatment of primary islets with a PAR2 agonist (2fLI) in combination with inhibitors of insulin secretion and signaling was sufficient to induce insulin and PAX4 gene expression. Moreover, in primary human islets, this treatment also led to the induction of bihormonal islet cells coexpressing glucagon and insulin, a hallmark of islet cell transdifferentiation. Mechanistically, insulin inhibited the positive effect of a PAR2 agonist on insulin gene expression and also led to an increase in PAX4, which plays an important role in islet cell transdifferentiation. The studies presented here demonstrate that insulin represses transdifferentiation of α- to β-cells induced by activation of PAR2. This provides a mechanistic explanation for the observation that α- to β-cell transdifferentiation occurs only in the setting of severe β-cell ablation. The mechanistic understanding of islet cell transdifferentiation and the ability to modulate that process using available pharmacological reagents represents an important step along the path towards harnessing this novel mechanism of β-cell neogenesis as a therapy for diabetes.

Generation and Phenotype Identification of PAX4 Gene Knockout Rabbit by CRISPR/Cas9 System.

Paired-homeodomain transcription factor 4 (PAX4) gene encodes a transcription factor which plays an important role in the generation, differentiation, development, and survival of insulin-producing β-cells during mammalian pancreas development. PAX4 is a key diabetes mellitus (DM) susceptibility gene, which is associated with many different types of DM, including T1DM, T2DM, maturity onset diabetes of the young 9 (MODY9) and ketosis prone diabetes. In this study, a novel PAX4 gene knockout (KO) model was generated through co-injection of clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9 (Cas9) mRNA/sgRNA into rabbit zygotes. Typical phenotypes of growth retardation, persistent hyperglycemia, decreased number of insulin-producing β cells and increased number of glucagon-producing α cells were observed in the homozygous PAX4 KO rabbits. Furthermore, DM associated phenotypes including diabetic nephropathy, hepatopathy, myopathy and cardiomyopathy were also observed in the homozygous PAX4 KO rabbits but not in the wild type (WT) controls and the heterozygous PAX4 KO rabbits. In summary, this is the first PAX4 gene KO rabbit model generated by CRISPR/Cas9 system. This novel rabbit model may provide a new platform for function study of PAX4 gene in rabbit and gene therapy of human DM in clinical trails.

Pax4 Gene Transfer Induces α-to-ß Transdifferentiation and Improves the Therapeutic Efficacy of Islet Transplantation

Pax4 is a transcriptional factor which plays a critical role in the generation of insulin producing β-cells during embryonic development. Our group previously demonstrated that Pax4 delivered by adenoviral vector induces insulin expression and reduces glucagon in αTC1.9 cells and primary human islets. The purpose of the present study was to determine whether Pax4 gene expression induces pancreatic α-cells to become β-cell using lineage tracing techniques and confers therapeutic benefits in the context of islet transplantation. In this study, we generated α mouse model in which their α cells are labelled with YFP, namely αYFP mouse. Using this mouse model, we found that Ad5.Pax4 induced glucagon secreting α-cells to become insulin secreting β-cells in isolated αYFP islets. Quantification of YFP+/Pax4+/insulin+ cells confirmed that Pax4 induced α-to-β cell conversion as assessed by the presence of YFP reporter in new β-cells. In addition, these Pax4-treated islets increased glucose-stimulated insulin secretion compare to control group. Furthermore, we transplanted Pax4-treated primary mouse islets into kidney capsule of the streptozotocin-induced diabetic mice and found that they significantly reduced the blood glucose and improved glucose tolerance compared to control groups. These data indicate that Pax4 induced α- to functional β-cell conversion, and proved therapeutic benefits for islet transplantation. Disclosure K.R. Parajuli: None. Y. Zhang: None. H. Wu: None.

The type 2 diabetes-associated HMG20A gene is mandatory for islet beta cell functional maturity

HMG20A (also known as iBRAF) is a chromatin factor involved in neuronal differentiation and maturation. Recently small nucleotide polymorphisms (SNPs) in the HMG20A gene have been linked to type 2 diabetes mellitus (T2DM) yet neither expression nor function of this T2DM candidate gene in islets is known. Herein we demonstrate that HMG20A is expressed in both human and mouse islets and that levels are decreased in islets of T2DM donors as compared to islets from non-diabetic donors. In vitro studies in mouse and human islets demonstrated that glucose transiently increased HMG20A transcript levels, a result also observed in islets of gestating mice. In contrast, HMG20A expression was not altered in islets from diet-induced obese and pre-diabetic mice. The T2DM-associated rs7119 SNP, located in the 3′ UTR of the HMG20A transcript reduced the luciferase activity of a reporter construct in the human beta 1.1E7 cell line. Depletion of Hmg20a in the rat INS-1E cell line resulted in decreased expression levels of its neuronal target gene NeuroD whereas Rest and Pax4 were increased. Chromatin immunoprecipitation confirmed the interaction of HMG20A with the Pax4 gene promoter. Expression levels of Mafa, Glucokinase, and Insulin were also inhibited. Furthermore, glucose-induced insulin secretion was blunted in HMG20A-depleted islets. In summary, our data demonstrate that HMG20A expression in islet is essential for metabolism-insulin secretion coupling via the coordinated regulation of key islet-enriched genes such as NeuroD and Mafa and that depletion induces expression of genes such as Pax4 and Rest implicated in beta cell de-differentiation. More importantly we assign to the T2DM-linked rs7119 SNP the functional consequence of reducing HMG20A expression likely translating to impaired beta cell mature function.

Association of PAX4 gene R192H polymorphisms in Chinese Han type 2 diabetes

The purpose of this study was to explore whether PAX4 gene R192H polymorphism plays a role in type 2 diabetes in Chinese Han population, using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technology to detect R192H polymorphism in exon 5 of PAX4 gene in 410 cases of patients with type 2 diabetes and 95 cases of non-diabetic controls from January 2014 to January 2015. There are no significant differences in the genotypes or allele frequencies between type 2 diabetes mellitus (T2DM) and non-diabetic controls about PAX4 R192H polymorphism. Similarly, no statistical differences were observed in the T2DM and non-diabetic controls when adjusting for gender stratification and age stratification. The comparisons on the subjects in clinical characteristics including blood glucose, glycated hemoglobin, cholesterol, triglycerides, high-density lipoprotein cholesterol, and systolic blood pressure showed no difference between carriers with the GA and AA genotype except diastolic blood pressure (p value = 0.038). PAX4 gene exon 5 R192H polymorphism was not associated with type 2 diabetes in Chinese Han population.

Investigation for PAX4 gene R192H mutation of a MODY family

Investigation for PAX4 gene R192H mutation of a MODY family

Monogenic diabetes associated with PAX4 gene mutations (MODY9): first description in Russia

Maturity-onset diabetes of the young (MODY) is a heterogeneous group of disorders characterised by autosomal dominant type of inheritance and caused by genetic defects leading to dysfunction of pancreatic beta-cells. To date, at least 13 subtypes of MODY have been described in the literature, the most frequent of which are MODY types 13. MODY2 and MODY3 are the most prevalent subtypes, and were previously described in our country, Russia. Several cases of rare MODY subtypes were subsequently described in the Russian literature. The current report is the first in the Russian literature to present clinical and molecular genetic characteristics of two cases of another rare MODY subtypeMODY9. This type of MODY is associated with mutations in the PAX4 gene, which encodes transcription factor PAX4, one of the factors essential for pancreatic beta-cell differentiation. Molecular genetic analysis was performed using next-generation sequencing, a new method recently applied to verify monogenic diseases and, in particular, MODY. This study reports a novel mutation in the PAX4 gene in MODY patients.

Improved insulin-secreting properties of pancreatic islet mesenchymal stem cells by constitutive expression of Pax4 and MafA.

For long-term treatment of diabetes type 1, transplantation of insulin-producing beta cells may be a promising method, but the limited number of islets for transplantation requires the development of different approaches. In this study, we aimed to generate betalike insulin-producing cells. For this purpose, MafA, Pax4, and Ngn3 genes were transferred into pancreatic islet-derived mesenchymal stem cells, and the effect of their ectopic expressions on differentiation efficiency was examined. Stemness properties of pancreatic islet stem cells were characterized. The 3 genes were transfected by electroporation and expressed constitutively. The transfected cells were further stimulated to differentiate by using chemical induction. Pax4 expression had significant effects on differentiation into insulin-producing cells. Although it caused morphological alterations in cells, similar to epithelial cells, the insulin secretion levels remained lower than those of the cell line cotransfected with MafA and Pax4. Cotransfection of the 3 transcription factors did not further improve the beta-like cell generation. MafA and Pax4 ectopic expression resulted in improved differentiation efficiency into insulin-secreting cells. However, support of this differentiation process using additional chemical induction may sufice to overcome control by endogenous regulatory pathways.

PAX4 Gene Transfer Induces α-to-β Cell Phenotypic Conversion and Confers Therapeutic Benefits for Diabetes Treatment.

The transcription factor Pax4 plays a critical role in the determination of α- versus β-cell lineage during endocrine pancreas development. In this study, we explored whether Pax4 gene transfer into α-cells could convert them into functional β-cells and thus provide therapeutic benefits for insulin-deficient diabetes. We found that Pax4 delivered by adenoviral vector, Ad5.Pax4, induced insulin expression and reduced glucagon expression in αTC1.9 cells. More importantly, these cells exhibited glucose-stimulated insulin secretion, a key feature of functional β-cells. When injected into streptozotocin-induced diabetic mice, Pax4-treated αTC1.9 cells significantly reduced blood glucose, and the mice showed better glucose tolerance, supporting that Pax4 gene transfer into αTC1.9 cells resulted in the formation of functional β-cells. Furthermore, treatment of primary human islets with Ad5.Pax4 resulted in significantly improved β-cell function. Detection of glucagon(+)/Pax4(+)/Insulin(+) cells argued for Pax4-induced α-to-β cell transitioning. This was further supported by quantification of glucagon and insulin bi-hormonal cells, which was significantly higher in Pax4-treated islets than in controls. Finally, direct administration of Ad5.Pax4 into the pancreas of insulin-deficient mice ameliorated hyperglycemia. Taken together, our data demonstrate that manipulating Pax4 gene expression represents a viable therapeutic strategy for the treatment of insulin deficient diabetes.

MTOR Inhibition: Reduced Insulin Secretion and Sensitivity in a Rat Model of Metabolic Syndrome.

Sirolimus (SRL) has been associated with new-onset diabetes mellitus after transplantation. The aim was to determine the effect of SRL on development of insulin resistance and β -cell toxicity. Lean Zucker rat (LZR) and obese Zucker rat (OZR) were distributed into groups: vehicle and SRL (0.25, 0.5, or 1.0 mg/kg) during 12 or 28 days. Intraperitoneal glucose tolerance test (IPGTT) was evaluated at days 0, 12, 28, and 45. Islet morphometry, β-cell proliferation, and apoptosis were analyzed at 12 days. Islets were isolated to analyze insulin content, insulin secretion, and gene expression. After 12 days, SRL treatment only impaired IPGTT in a dose-dependent manner in OZR. Treatment prolongation induced increase of area under the curve of IPGTT in LZR and OZR; however, in contrast to OZR, LZR normalized glucose levels after 2 hours. The SRL reduced pancreas weight and islet proliferation in LZR and OZR as well as insulin content. Insulin secretion was only affected in OZR. Islets from OZR + SRL rats presented a downregulation of Neurod1, Pax4, and Ins2 gene. Genes related with insulin secretion remained unchanged or upregulated. In conditions that require adaptive β -cell proliferation, SRL might reveal harmful effects by blocking β -cell proliferation, insulin production and secretion. These effects disappeared when removing the therapy.