Research on human embryos and cloning : difficulties of legislating in a changing environment and model approaches to regulation

It is difficult to regulate rapidly changing fields of science. New technologies are not anticipated and legislation becomes inadequate. Legislative definitions are also problematic. This article begins with consideration of such difficulties in the context of research on human embryos and cloning. It considers problems with past legislative definitions in Australia, the new regulatory regime, and whether that regime now sets clear boundaries. It is found that problems still exist – some terms are not adequately defined and boundaries for research prove unclear. Three regulatory approaches are therefore discussed. Legislation based on strict definitions is compared to a legislative model that leaves terms undefined. The third model – which combines framework legislation with the oversight of a regulatory authority – is seen as most suitable. However, problems with this model are recognised and suggestions made regarding how to ensure the “framework” remains workable and effective.

Regulatory design strategies and enforcement approaches for research involving human embryos and cloning in Australia and the United Kingdom – time for a change.

This paper examines regulatory design strategies and enforcement approaches in the context of the UK and Australia’s regulation of research involving human embryos and cloning. The aim is to discuss current regulation in view of the impending review of the Research Involving Human Embryos Act 2002 (Cth) and the Prohibition of Human Reproductive Cloning Act 2002 (Cth). It is argued that the type of regulation used in relation to those who are licensed to research in Australia is unsuitable due to an over-emphasis on deterrence and the authoritarian approach taken by regulatory bureaucracies. The cost and efficiency of the current system is also questioned. The central thesis is that a co-regulatory system that combines the existing framework legislation with self-regulation should be adopted for licence holders. Such regulation of licence holders should include responsive regulatory strategies. ‘Command and control’ design strategies and deterrence approaches present in the current regulatory systems for breaches of legislation by non-licence holders and serious breaches by licence holders should be maintained.

A novel zebrafish jak2a V581F model shared features of human JAK2 V617F polycythemia vera

Objective : The Janus kinase 2 (JAK2) is important for embryonic primitive hematopoiesis. A gain-of-function JAK2 (JAK2^V617F) mutation in human is pathogenetically linked to polycythemia vera (PV). In this study, we generated a zebrafish ortholog of human JAK2^V617F (referred herewith jak2a^V581F) by site-directed mutagenesis and examined its relevance as a model of human PV.

Materials and Methods : Zebrafish embryos at one-cell stage were injected with jak2a^V581F mRNA (200pg/embryo). In some experiments, the embryos were treated with a specific JAK2 inhibitor, TG101209. The effects of jak2a stimulation on hematopoiesis, jak/stat signaling, and erythropoietin signaling were evaluated at 18-somites.

Results : Injection with jak2a^V581F mRNA significantly increased erythropoiesis, as enumerated by flow cytometry based on gfp+ population in dissociated Tg(gata1:gfp) embryos. The response was reduced by stat5.1 morpholino coinjection (control: 4.37% ± 0.08%; jak2a^V581F injected: 5.71% ± 0.07%, coinjecting jak2a^V581F mRNA and stat5.1 morpholino: 4.66% ± 0.13%; p < 0.01). jak2a^V581F mRNA also upregulated gata1 (1.83 ± 0.08 fold; p = 0.005), embryonic α-hemoglobin (1.61 ± 0.12 fold; p = 0.049), and β-hemoglobin gene expression (1.65 ± 0.13–fold; p = 0.026) and increased stat5 phosphorylation. These responses were also ameliorated by stat5.1 morpholino coinjection or treatment with a specific JAK2 inhibitor, TG101209. jak2a^V581F mRNA significantly reduced erythropoietin gene (0.24 ± 0.03 fold; p = 0.006) and protein expression (control: 0.633 ± 0.11; jak2a^V581F mRNA: 0.222 ± 0.07 mIU/mL; p = 0.019).

Conclusion : The zebrafish jak2a^V581F model shared many features with human PV and might provide us with mechanistic insights of this disease.

The small molecule, genistein, increases hepcidin expression in human hepatocytes

Hepcidin, a peptide hormone that decreases intestinal iron absorption and macrophage iron release, is a potential drug target for patients with iron overload syndromes because its levels are inappropriately low in these individuals. Endogenous stimulants of Hepcidin transcription include bone morphogenic protein 6 (BMP6) and interleukin-6 (IL-6) by effects on mothers against decapentaplegic homolog (Smad)4 or signal transducer and activator of transcription (Stat)3, respectively. We conducted a small-scale chemical screen in zebrafish embryos to identify small molecules that modulate hepcidin expression. We found that treatment with the isoflavone, genistein, from 28-52 hours postfertilization in zebrafish embryos enhanced Hepcidin transcript levels, as assessed by whole-mount in situ hybridization and quantitative real-time reverse-transcriptase polymerase chain reaction. Genistein's stimulatory effect was conserved in human hepatocytes: Genistein treatment of HepG2 cells increased both Hepcidin transcript levels and promoter activity. We found that genistein's effect on Hepcidin expression did not depend on estrogen receptor signaling or increased cellular iron uptake, but was impaired by mutation of either BMP response elements or the Stat3-binding site in the Hepcidin promoter. RNA sequencing of transcripts from genistein-treated hepatocytes indicated that genistein up-regulated 68% of the transcripts that were up-regulated by BMP6; however, genistein raised levels of several transcripts involved in Stat3 signaling that were not up-regulated by BMP6. Chromatin immunoprecipitation and ELISA experiments revealed that genistein enhanced Stat3 binding to the Hepcidin promoter and increased phosphorylation of Stat3 in HepG2 cells. Conclusion: Genistein is the first small-molecule experimental drug that stimulates Hepcidin expression in vivo and in vitro. These experiments demonstrate the feasibility of identifying and characterizing small molecules that increase Hepcidin expression. Genistein and other candidate molecules may subsequently be developed into new therapies for iron overload syndromes.