Assays to Detect β-Tubulin Codon 200 Polymorphism in Trichuris trichiura and Ascaris lumbricoides

The soil-transmitted helminths Ascaris lumbricoides and Trichuris trichiura are gastrointestinal nematodes causing many disabilities in tropical parts of the developing world. Control programs, such as “The Focussing Resources on Effective School Health” (FRESH) Partnership, have been implemented to remove human soil-transmitted nematodes through large-scale use of benzimidazole anthelmintic drugs for school-aged children in developing countries. The benzimidazole drugs albendazole and mebendazole are commonly used as a single annual treatment in areas where the burden is high. In veterinary nematodes, repeated use of these anthelmintics has selected for resistant populations. Resistance to benzimidazoles is commonly associated with a single amino acid substitution from phenylalanine to tyrosine in the β-tubulin gene at position 200. In this study, we have developed pyrosequencing assays for codon 200 in A. lumbricoides and T. trichiura to screen for this single nucleotide polymorphism (SNP) in β-tubulin. The 200Tyr SNP was detected at low frequency in T. trichiura from non-treated people from Kenya and at high frequency in T. trichiura from treated people from Panama. The presence of the resistance-associated SNP may play a role in the sometimes low and variable efficacy of benzimidazole anthelmintics against T. trichiura

Lrp5 Is Not Required for the Proliferative Response of Osteoblasts to Strain but Regulates Proliferation and Apoptosis in a Cell Autonomous Manner

Although Lrp5 is known to be an important contributor to the mechanisms regulating bone mass, its precise role remains unclear. The aim of this study was to establish whether mutations in Lrp5 are associated with differences in the growth and/or apoptosis of osteoblast-like cells and their proliferative response to mechanical strain in vitro. Primary osteoblast-like cells were derived from cortical bone of adult mice lacking functional Lrp5 (Lrp5−/−), those heterozygous for the human G171V High Bone Mass (HBM) mutation (LRP5G171V) and their WT littermates (WTLrp5, WTHBM). Osteoblast proliferation over time was significantly higher in cultures of cells from LRP5G171V mice compared to their WTHBM littermates, and lower in Lrp5−/− cells. Cells from female LRP5G171V mice grew more rapidly than those from males, whereas cells from female Lrp5−/− mice grew more slowly than those from males. Apoptosis induced by serum withdrawal was significantly higher in cultures from Lrp5−/− mice than in those from WTHBM or LRP5G171V mice. Exposure to a single short period of dynamic mechanical strain was associated with a significant increase in cell number but this response was unaffected by genotype which also did not change the ‘threshold’ at which cells responded to strain. In conclusion, the data presented here suggest that Lrp5 loss and gain of function mutations result in cell-autonomous alterations in osteoblast proliferation and apoptosis but do not alter the proliferative response of osteoblasts to mechanical strain in vitro

Strain uses gap junctions to reverse stimulation of osteoblast proliferation by osteocytes (vol 35, pg 56, 2017)

Biotechnology and Biological Sciences Research Council BB/I014608/1, Arthritis Research UK 20413, 20581, 20039