Lack of Wdr13 Gene in Mice Leads to Enhanced Pancreatic Beta Cell Proliferation, Hyperinsulinemia and Mild Obesity

WD-repeat proteins are very diverse, yet these are structurally related proteins that participate in a wide range of cellular functions. WDR13, a member of this family, is conserved from fishes to humans and localizes into the nucleus. To understand the in vivo function(s) of Wdr13 gene, we have created and characterized a mutant mouse strain lacking this gene. The mutant mice had higher serum insulin levels and increased pancreatic islet mass as a result of enhanced beta cell proliferation. While a known cell cycle inhibitor, p21, was downregulated in the mutant islets, over expression of WDR13 in the pancreatic beta cell line (MIN6) resulted in upregulation of p21, accompanied by retardation of cell proliferation. We suggest that WDR13 is a novel negative regulator of the pancreatic beta cell proliferation. Given the higher insulin levels and better glucose clearance in Wdr13 gene deficient mice, we propose that this protein may be a potential candidate drug target for ameliorating impaired glucose metabolism in diabetes

Anorectal agenesis with rectovaginal fistula: A rare/regional variant

Aims and Objectives: The anatomical types of female anorectal malformation (ARM) are complex and include several rare and regional variants. The aim of this report is to highlight the diagnosis and management of cases of anorectal agenesis with rectovaginal fistula (RVF). Materials and Methods: This is a retrospective chart review of cases of RVF who underwent definitive surgery between November 2000 and September 2016 in a single institution. Results: Of the 244 female patients with anorectal anomalies treated in our institution over a 16-year period, there were 15 cases of RVF with anorectal agenesis giving it an incidence of 6%. All cases of RVF with anorectal agenesis presented with absence of anal opening, varying degree of abdominal distension, and history of passage of fecal matter per vaginum. The mean age of presentation was 3 months (newborn to 1 year). Sigmoid colostomy was done in 13 cases. Distal colostogram demonstrated high RVF which was further confirmed by examination under anesthesia (EUA) and endoscopy before definitive surgery. With the posterior sagittal approach, division of the RVF and anorectoplasty was possible in 14 cases; however, one patient required an abdominal mobilization of the colon for pull through due to a very short distal bowel segment. During a mean postoperative follow-up of 5 years, cosmetic and functional results were satisfactory. Conclusion: RVF with anorectal agenesis is a rare/regional variant of female ARMs. Clinical examination along with distal colostogram, EUA, and endoscopy clinches the diagnosis. Anorectal reconstruction by posterior sagittal anorectoplasty results in a satisfactory outcome

Pediatric esophageal substitution by gastric pull-up and gastric tube

Aim: The aim of this study was to report the results of pediatric esophageal substitution by gastric pull-up (GPU) and gastric tube (GT) from a tertiary care pediatric center. Materials and Methods: Retrospective analysis of the surgical techniques, results, complications, and final outcome of all pediatric patients who underwent esophageal substitution in a single institution was performed. Results: Twenty-four esophageal substitutions were performed over 15-year period. The indications were pure esophageal atresia (EA)-19, EA with distal trachea-esophageal fistula-2, EA with proximal pouch fistula-1, and esophageal stricture in two patients. Mean age and weight at operation were 17 months and 9.5 kg, respectively. GPU was the most common procedure (19) followed by reverse GT (4) and gastric fundal tube (1). Posterior mediastinal and retrosternal routes were used in 17 and 7 cases, respectively. Major complications included three deaths in GPU cases resulting from postoperative tachyarrhythmias leading to cardiac arrest, cervical anastomotic leak-17, and anastomotic stricture in six cases. Perioperative tachyarrhythmias (10/19) and transient hypertension (2/19) were observed in GPU patients, and they were managed with beta blocker drugs. Postoperative ventilation in Intensive Care Unit was performed for all GPU, but none of the GT patients. Follow-up ranged from 6 months to 15 years that showed short-term feeding difficulties and no major growth-related problems. Conclusions: Perioperative tachyarrhythmias are common following GPU which mandates close intensive care monitoring with ventilation and judicious use of beta blocking drugs. Retrosternal GT with a staged neck anastomosis can be performed without postoperative ventilation

Glucose tolerance test (GTT) and Insulin tolerance test (ITT) of <i>Wdr13</i> knockout mice.

<p>A) GTT at 2 months on normal chow (n = 8–12). B) GTT at 12 months on normal chow (n = 8–10). C) Glucose tolerance test at 6 months on high fat diet (n = 8). D) ITT at 2 months on normal chow (n = 8–12). E) ITT at 12 months on normal chow (n = 8–10). F) Insulin tolerance test at 6 months on high fat diet (n = 8) G) ITT at 9 months on high fat diet (n = 6–8). (+/0 wild type male; −/0 knockout male).</p

Generation of <i>Wdr</i>13 knockout mice.

<p>A) Exon2 and exon3 (partial) of <i>Wdr</i>13 gene were replaced by neomycin resistance marker gene. B) Southern blot analysis showing 9.3 kb <i>Eco</i>RI fragment from the wild type allele and 3.8 kb fragment from the mutant allele using a 700 bp probe from 5′ end of the locus. C) Northern blot analysis by <i>Wdr</i>13 cDNA probe showing the absence of <i>Wdr</i>13 transcript in <i>Wdr</i>13−/0 mice (upper panel) and ethidium bromide staining as loading control (lower panel). D) Western blot analysis from testis, pancreas and purified islets using anti-WDR13 rabbit polyclonal antibody (upper panel) showing the absence of WDR13 protein in <i>Wdr</i>13−/0 mice, and anti-beta actin as loading control (lower panel). The expression of WDR13 is extremely high in islets in comparison to acinar cells. E) Expression of WDR13 protein in insulin sensitive tissues such as liver, pancreas, muscle and adipose tissues using anti WDR13 antibody (upper panel) and posceau stained blot as loading control (lower panel). (+/0 wild type male; −/0 knockout male).</p

Histology of pancreatic islet, pancreatic beta cell proliferation, in-vivo and in-vitro insulin secretion in <i>Wdr</i>13 knockout mice.

<p>A) Islet morphology showing increased islet mass by H&E staining. B) Islet mass in mg at 6 months on high fat diet showing increased total islet mass in <i>Wdr</i>13 knockout mice (n = 4). C) Percent of dividing cells in islets by BrdU labeling (n = 309–517). D) Analysis of pancreatic beta cells proliferation analysis by BrdU labeling showing more proliferating beta cells in <i>Wdr13</i> knockout islets. E) In-vivo insulin secretion in response to glucose at 6 months (n = 4 to 6). F) In-vitro insulin secretion from isolated pancreatic islet of the wild type and <i>Wdr</i>13 knockout mice at 2.8 mM or 16 mM glucose concentration (+/0 wild type male; −/0 knockout male).</p

A draft genome sequence of the pulse crop chickpea (Cicer arietinum L.)

Cicer arietinum L. (chickpea) is the third most important food legume crop. We have generated the draft sequence of a desi-type chickpea genome using next-generation sequencing platforms, bacterial artificial chromosome end sequences and a genetic map. The 520-Mb assembly covers 70% of the predicted 740-Mb genome length, and more than 80% of the gene space. Genome analysis predicts the presence of 27 571 genes and 210 Mb as repeat elements. The gene expression analysis performed using 274 million RNA-Seq reads identified several tissue-specific and stress-responsive genes. Although segmental duplicated blocks are observed, the chickpea genome does not exhibit any indication of recent whole-genome duplication. Nucleotide diversity analysis provides an assessment of a narrow genetic base within the chickpea cultivars. We have developed a resource for genetic markers by comparing the genome sequences of one wild and three cultivated chickpea genotypes. The draft genome sequence is expected to facilitate genetic enhancement and breeding to develop improved chickpea varieties