Male Fertility Defect Associated with Disrupted BRCA1-PALB2 Interaction in Mice

PALB2 links BRCA1 and BRCA2 in homologous recombinational repair of DNA double strand breaks (DSBs). Mono-allelic mutations in PALB2 increase the risk of breast, pancreatic, and other cancers, and biallelic mutations cause Fanconi anemia (FA). Like Brca1 and Brca2, systemic knock-out of Palb2 in mice results in embryonic lethality. In this study, we generated a hypomorphic Palb2 allele expressing a mutant PALB2 protein unable to bind BRCA1. Consistent with an FA-like phenotype, cells from the mutant mice showed hypersensitivity and chromosomal breakage when treated with mitomycin C, a DNA interstrand crosslinker. Moreover, mutant males showed reduced fertility due to impaired meiosis and increased apoptosis in germ cells. Interestingly, mutant meiocytes showed a significant defect in sex chromosome synapsis, which likely contributed to the germ cell loss and fertility defect. Our results underscore the in vivo importance of the PALB2-BRCA1 complex formation in DSB repair and male meiosis.     

Ret receptor tyrosine kinase sustains proliferation and tissue maturation in intestinal epithelia

Expression of the Ret receptor tyrosine kinase is a defining feature of enteric neurons. Its importance is underscored by the effects of its mutation in Hirschsprung disease, leading to absence of gut innervation and severe gastrointestinal symptoms. We report a new and physiologically significant site of Ret expression in the intestine: the intestinal epithelium. Experiments in Drosophila indicate that Ret is expressed both by enteric neurons and adult intestinal epithelial progenitors, which require Ret to sustain their proliferation. Mechanistically, Ret is engaged in a positive feedback loop with Wnt/Wingless signalling, modulated by Src and Fak kinases. We find that Ret is also expressed by the developing intestinal epithelium of mice, where its expression is maintained into the adult stage in a subset of enteroendocrine/enterochromaffin cells. Mouse organoid experiments point to an intrinsic role for Ret in promoting epithelial maturation and regulating Wnt signalling. Our findings reveal evolutionary conservation of the positive Ret/Wnt signalling feedback in both developmental and homeostatic contexts. They also suggest an epithelial contribution to Ret loss‐of‐function disorders such as Hirschsprung disease.     

The inadequate smear: does it matter?

The objective of this study was to quantify the incidence of underlying cervical intraepithelial neoplasia (CIN) among women referred for colposcopy with three consecutive inadequate smears. The design was a retrospective cohort study analysing data from a regional colposcopy database at Cervical Screening Wales. Women who were referred to all the colposcopy clinics in Wales with three consecutive inadequate smears, the third inadequate smear being taken between 1 April 2001 and 31 March 2002 constituted the study population. The results of the fourth smear taken at the colposcopy clinic after three consecutive inadequate smears, the worst biopsy results from any of the subsequent colposcopies and the relationship between the result of the fourth smear taken at colposcopy clinic and any histology result were the main outcome measures. The accuracy of the colposcopic opinion was also examined. Of the 433 women identified as having been referred because of three consecutive inadequate smears, 302 were linked to either a subsequent smear and/or a biopsy result. An adequate smear result was available for 85% of these women when the smear was taken in the colposcopy clinic; 77% were reported as negative and 8% were abnormal. Of the 347 women seen in the colposcopy clinic, high-grade CIN was seen in 3% and low-grade lesion in 8%. The sensitivity and specificity of the fourth inadequate smear test in predicting underlying CIN were 15% and 84% respectively, with a positive predictive value of 8%. The sensitivity and specificity of colposcopy in predicting histological CIN among patients with three inadequate smears was 70% and 49%, respectively, and the positive predictive value was 44%. This study raises the question as to whether three consecutive inadequate smears should be considered as an indication for colposcopy, or merely for a further smear to be taken in circumstances where there is a greater likelihood getting an adequate result.     

Preservation of polyclonal antibody production by hybridization

The fusion of unimmunized (Balb/c × SJL)_F1, mouse spleen cells in which a polyclonal response had been induced by bacterial lipopolysaccharide with a myeloma cell line resulted in hybrid cell populations. The hybrid populations obtained elaborated antibody activity to human hemoglobin A, Keyhole Limpet hemocyanin, the dinitrophenyl (DNP) hapten, and human erythrocytes, Thus, hybridization allowed preservation of the normally transitory polyclonal response induced in mouse B cells by lipopolysaccharide. Furthermore, monospecific production of anti-DNP antibody was successfully factored out of the polyspecific production of antibodies by cloning. The expansion and subsequent injection of one of these clones into a (Balb/c × SJL)_F1 mouse resulted in the formation of an antibody-producing tumor that was successfully passed to other (Balb/c × SJL)_F1 recipients. Collection of the serum from tumor-bearing mice provided useful quantities of an anti-DNP antiserum without resort to any program of immunization whatsoever.