Phosphatase-defective LEOPARD syndrome mutations in PTPN11 gene have gain-of-function effects during Drosophila development

Missense mutations in the PTPN11 gene, which encodes the proteintyrosine phosphatase SHP-2, cause clinically similar but distinctivedisorders, LEOPARD (LS) and Noonan (NS) syndromes. The LS isan autosomal dominant disorder with pleomorphic developmentalabnormalities including lentigines, cardiac defects, short statureand deafness. Biochemical analyses indicated that LS allelesengender loss-of-function (LOF) effects, while NS mutationsresult in gain-of-function (GOF). These biochemical findingslead to an enigma that how PTPN11 mutations with opposite effectson function result in disorders that are so similar. To studythe developmental effects of the commonest LS PTPN11 alleles(Y279C and T468M), we generated LS transgenic fruitflies usingcorkscrew (csw), the Drosophila orthologue of PTPN11. Ubiquitousexpression of the LS csw mutant alleles resulted in ectopicwing veins and, for the Y279C allele, rough eyes with increasedR7 photoreceptor numbers. These were GOF phenotypes mediatedby increased RAS/MAPK signaling and requiring the LS mutant’sresidual phosphatase activity. Our findings provide the firstevidence that LS mutant alleles have GOF developmental effectsdespite reduced phosphatase activity, providing a rationalefor how PTPN11 mutations with GOF and LOF produce similar butdistinctive syndromes.     

Role of Ocimum sanctum as a Genoprotective Agent on Chlorpyrifos-Induced Genotoxicity

Protective effect of Ocimum sanctum was evaluated on chlorpyrifos-induced genotoxicity in in vivo and in vitro models. Two different concentrations of pesticide were taken, i.e., 1/5 and 1/15 of LD(50) of chlorpyrifos for the in vivo study. Rats were pre-treated orally with O. sanctum extract (OE) at 50 mg/kg b.wt. For the in vitro studies, human lymphocyte cultures were exposed to 75 μg/ml chlorpyrifos with and without OE. Structural and numerical (both aneuploidy and euploidy types) chromosomal aberrations (CAs) were scored for the assessment of induced genotoxic effects, while the variation in mitotic index (MI) was considered as a monitor for induced cellular toxicity. The same concentration of the pesticide (75 μg/ml) was taken to study the DNA damage by comet assay. Results showed that lymphocytes treated with the pesticide exhibited increased DNA damage but the increase was statistically insignificant (P>0.05). In rats pretreated with OE, a significant (P<0.01) increase in MI was observed and there was a significant decrease in the frequency of aberrant cells as compared to the rats treated with chlorpyrifos alone. A significant (P<0.05) increase in CA was observed in cultures treated with 75 μg/ml chlorpyrifos as compared to controls, which decreased significantly (P<0.05) with OE pretreatment.     

Paramedic-conducted Mental Health Counselling for Abused Women in Rural Bangladesh: An Evaluation from the Perspective of Participants

This paper reports on evaluation of an initiative to use paramedics as the first-level mental health counsellors of abused women in rural Bangladesh (2003–2004) from the perspective of the abused women who participated in one or more counselling sessions. Thirty in-depth interviews, followed by a survey (n=372), targeted to cover all participants, were conducted in 2006. Overall, the arrangement, management of ethical issues, and skills of paramedics were rated favourably. Most (89%) abused women (n=372) considered the session useful; one-fourth of these women considered it very useful; and only a few abused women considered the session useless. Usefulness of the session was expressed mostly in terms of relief attained after talking about the issue. Most (87%) women reported being encouraged to be self-confident. In a context characterized by low self-confidence of women, lack of opportunity to talk about violence, and absence of professional mental health counselling services, this initiative is sufficiently promising to warrant further testing.Key words: Battered women, Counselling, Mental health, Paramedics, Bangladesh     

Nodular amiodarone lung disease

The antiarrhythmic drug amiodarone accumulates in many organs of the body. Amiodarone lung disease (ALD) most commonly manifests clinically as an interstitial pneumonitis. The few reports of nodular ALD generally have been in the clinical and radiographic literature. No detailed histopathologic analysis of nodular ALD is available. We report 4 patients with nodular ALD, all of whom had excision of a nodule and none had a preoperative diagnosis of ALD. The radiographic suspicion before excisional biopsy in all 4 cases was malignancy. The initial pathologic suspicion in all 4 cases was either an abscess or vasculitis. In 3 of the 4 cases, where the dosages were known, each patient received 800 mg/d for 7 or more months. All cases have strikingly similar histopathology, with vacuolated histiocytes massed within alveoli to form macroscopic nodules with tissue breakdown. Ultrastructural examination of lung and peripheral nerve in 1 case showed the characteristic inclusions of amiodarone in the cytoplasm of swollen histiocytes. Vacuolated histiocytes filled with these inclusions indicate the effect of the drug, but in these 4 cases there was tissue destruction, which indicated disease. The necrotizing nature of the massed histiocytes in the absence of infection or obstruction suggests the correct etiologic diagnosis even when the history of administration of the drug is not available at initial review.     

Laparoscopic correction of biliary duct stenosis in choledochal cyst

Background/Purposes

Laparoscopy has been widely accepted as a technique for the excision of choledochal cyst, but there has been little experience using it as a therapeutic modality for hepatic duct stenosis. The aim of this study is to present our experiences in laparoscopic excision of biliary stenosis and Roux-en-Y reconstruction for patients with choledochal cysts.

Methods

Eight patients, 3 boys and 5 girls (ranged from 6 months to 12 years; median age, 3.6 years), with hepatic duct stenosis underwent laparoscopic excision of the cyst and ductoplasty, with a Roux-en-Y hepaticoenterostomy between July 2001 and January 2005. Seven of the 8 patients had common hepatic duct stenosis with intrahepatic duct dilatation, and 1 had right hepatic duct stenosis with proximal dilatation.Four ports were inserted for instruments of 3- and 5-mm sizes. Each patient underwent a laparoscopic cholangiography. The gallbladder and dilated bile ducts were completely excised. The strictures of the hepatic ducts were treated by ductoplasty. The cut end of the duct was widened by incising along the anterior wall of the hepatic duct after excision of the narrow segment. By using intraoperative bile duct endoscopy, the optimal level of resection of the common hepatic duct was determined safely without endangering the orifices of the hepatic ducts or leaving any redundant duct, and the stone debris in the bile duct was identified and washed out. The Roux-en-Y jejunal loop was fashioned extracorporeally by exteriorizing the jejunum through the umbilical incision (1.0-1.5 cm) and passed up retrocolically followed by an end-to-side hepaticojejunostomy.

Results

The median duration of the operation was 4.3 hours (3.8-5.6 hours). Intraoperative bleeding was minimal, with no patients requiring blood transfusion. In 7 of the 8 cases, postoperative hospital stay ranged from 4 to 6 days; the other case had bile leak, which was cured by draining for 26 days without surgical intervention. All the patients had been followed up for 6 to 51 months. They stayed asymptomatic and well with no delayed complication.

Conclusions

Laparoscopically assisted hepatic ductoplasty is effective and safe for children with choledochal cyst. Bile duct endoscopy proved to be a valuable instrument in showing detailed variations of the biliary system and allowed a safe hepatic hilum exploration and accurate placed hepaticojejunal anastomosis.     

Recovery of liver perfusion after focal outflow obstruction and liver resection

BACKGROUND: Live liver donation requires extended liver resection in the donor with transection of the middle hepatic vein. This leads to focal outflow obstruction in the remnant liver or the partial graft. This study was designed to characterize the pathophysiological correlate of focal outflow obstruction in a small-for-size liver and its course of recovery in a rat model. METHODS: Ligation of the right median hepatic vein was combined with 50% hepatectomy. Microcirculation was visualized by orthogonal polarization spectroscopy after each operative step and before killing on days 1, 2, and 7. Histologic evaluation included morphological assessment, immunohistochemical determination of proliferation using BrdU, and laminin and von Willebrand factor expression, which both indicate vascularization of sinusoids. RESULTS: After ligation of the right median hepatic vein, congestion was visible and no sinusoidal blood flow was detected in the obstruction zone. By day 1 confluent centrilobular necrosis developed. Sinusoidal perfusion in the obstruction zone recovered partially. Many dilated vascularized sinusoidal canals connecting the obstruction zone with the normal zone were visible. Proliferative activity in the obstruction zone was markedly reduced compared with the normal zone. By day 7, liver parenchyma in the obstruction zone looked normal as did sinusoidal perfusion. In the border zone, few dilated vascular canals were apparent. CONCLUSION: Confluent centrilobular necrosis in the early postoperative phase, resulting from focal outflow obstruction, may be crucial for the development of a small-for-size syndrome. The exclusion of the outflow-obstructed zone from the functional liver mass during preoperative radiological risk assessment seems to be the logical consequence. Recovery of focal outflow obstruction occurs spontaneously by means of dilated sinusoids in the border zone, forming vascularized sinusoidal canals, which could serve as intrahepatic anastomosis.     

The Influence of Surface Roughness on the Displacement of Osteogenic Bone Particles during Placement of Titanium Screw‐Type Implants

Background: Previously, we demonstrated that bone debris, which is translocated during dental implant placement, has osteogenic potential. Therefore, it was hypothesized that implant surface roughness can influence the amount of translocated bone debris/particles and thereby the osteogenic response. Material and Methods: Small titanium implants were left turned (smooth) or blasted and acid etched. The implants were placed in fresh cadaver bone. After explantation, the implants were incubated in a culture medium containing β‐glycerophosphate and dexamethasone up to 24 days. Subsequently, histology, scanning electron microscopy (SEM), DNA analysis, and calcium (Ca) content measurements were performed. Results: For both types of implant during implant placement, bone particles were translocated because of inherent roughness of the implant. SEM and histology confirmed the presence of a bone‐like tissue on the surface of both types of implants, as also confirmed by DNA and Ca measurements. However, the significantly higher roughness of the etched implants accounted for more bone debris and accordingly elevated osteogenic response. Control samples, which had not been placed into bone, did not show mineralization in the same medium. Conclusion: The present study, for the first time, demonstrated that implant surface roughness can increase the amount of the translocated bone particles and thereby also have a beneficial effect on the osteogenic response of these bone particles. It is hypothesized that these bone fragments behave like miniature auto‐grafts and thereby play a significant role to enhance peri‐implant osteogenesis. Optimization of surface topography should be evaluated to take advantage of this additional effect of surface roughness.     

Evaluation of cytotoxic potential of structurally well-characterized RNA targeted ionic non-steroidal anti-inflammatory (NSAID) Cu(ii) & Zn(ii) DACH–mefenamato drug conjugates against human cancer cell lines

New RNA targeted ionic [Cu(DACH)₂(H₂O)₂](mef)₂, 1 and [Zn(DACH)₂(H₂O)₂](mef)₂, 2 drug conjugates were synthesized and characterized by spectroscopic techniques FT-IR, UV-vis, EPR in case of 1 and ¹H and ¹³C NMR in case of 2, ESI-MS, thermogravimetric analysis and single-crystal X-ray structure determination in case of 1. The interaction studies of 1 & 2 with most likely drug targets like ctDNA and tRNA were performed which demonstrated that the complexes 1 and 2 exhibited strong preferential binding to tRNA as compared to ctDNA, K_b = 2.52(±0.04) × 10⁵ M⁻¹, 7.85(±0.02) × 10⁴ M⁻¹, respectively. Scanning electron microscopy analyses of complex-ctDNA/tRNA condensates suggested the interaction of complexes with ctDNA/tRNA had occurred, followed by lengthening of DNA double helix and bulge region of tRNA. Cytotoxic activity of 1 and 2 against human cancer cell lines namely; MCF-7 (breast), HeLa (cervical), MIA-PA-CA 2 (pancreatic), A-498 (kidney), Hep-G2 (hepatoma) was evaluated by SRB assay. The obtained results showed that copper complex 1 was an outstanding cytotoxic agent with remarkably good GI₅₀ value (<10 μg ml⁻¹) against the tested cancer cell lines except for MIA-PA-CA 2, while zinc complex 2 revealed moderate cytotoxicity against all the tested cancer cell lines.

New RNA targeted ionic NSAID drug conjugates were evaluated for their antiproliferative activity against human cancer cell lines.