Abstract Gut bacterial β-glucuronidase (GUS) can reactivate xenobiotics that exert enterohepatic circulation- triggered gastrointestinal tract toxicity. GUS inhibitors can alleviate drug-induced enteropathy and improve treatment outcomes. We evaluated the inhibitory effect of Polygonum cuspidatum Siebold & Zucc. and its major constituents against Escherichia coli GUS (EcGUS), and characterized the inhibitory mechanism of each of the components. Trans-resveratrol 4’-O-β-D-glucopyranoside (HZ-1) and (-)-epicatechin gallate (HZ-2) isolated from P. cuspidatum were identified as the key components and potent inhibitors. These two components displayed strong to moderate inhibitory effects on EcGUS, with Ki values of 9.95 and 1.95 μM, respectively. Results from molecular docking indicated that HZ-1 and HZ-2 could interact with the key residues Asp163, Ser360, Ile 363, Glu413, Glu504, and Lys 568 of EcGUS via hydrogen bonding. Our findings demonstrate the inhibitory effect of P. cuspidatum and its two components on EcGUS, which supported the further evaluation and development of P. cuspidatum and its two active components as novel candidates for alleviating drug-induced damage in the mammalian gut.

RESUMO: O objetivo principal deste trabalho foi avaliar o impacto do programa de treinamento da escola de campo para agricultores (FFS) quanto ao nível de conhecimento dos agricultores (produtores de citros) em relação às melhores práticas de produção de citros. O Distrito de Sargodha foi a área de pesquisa escolhida para este estudo por ser o principal produtor de citros no Paquistão. Os dados quantitativos foram coletados por meio de entrevistas e analisados por meio do Statistical Package for Social Sciences (SPSS). Conclui-se que os jovens eram mais tendenciosos à obtenção de treinamento através do programa FFS do que os agricultores mais velhos. Os agricultores escolarizados estavam, em sua maioria, na categoria de agricultores FFS, enquanto que os agricultores analfabetos estavam, na categoria de não FF. Na categoria de FFS, a pecuária se mostrou o principal rendimento, conforme relatado pela maioria dos agricultores (31,3%), enquanto que, em 28% dos agricultores não-FFs, a principal fonte de renda era a venda de suas colheitas. Além disso, tanto os agricultores FFS como não FFS também ganham renda com fontes não agrícolas. O programa de treinamento FFS teve impacto positivo e significativo sobre o nível de conhecimento dos agricultores em relação às práticas de produção e proteção de citrus. O teste-T mostrou diferença altamente significativa (P<0,05) no nível de conhecimento dos agricultores FFS dos não-FFS.

ABSTRACT: The major objective of this paper was to assess the impact of the Farmer field school (FFS) training program on the knowledge level of farmers (citrus growers) regarding improved citrus production practices. Distract Sargodha was the targeted research area for this study being the leading producer of citrus in Pakistan. Quantitative data were collected through interview schedule and analyzed through Statistical Package for Social Sciences (SPSS). It was conclude that young people were more tend towards getting training under FFS program than old age farmers. Educated farmers were in majority in FFS farmers while; conversely illiterate farmers were in majority in the category of non-FFs farmers. In category of FFS farmers livestock farming was their primary income as reported by majority (31.3%) of FFS farmers and in the case of non-FFS farmer’s crop sale was their primary income source as reported by 28.4% of non-FFs farmers. In addition to that both FFS and non-FFS farmers also earn income from nonfarm sources like business, job, foreign remittances and labour. FFS training program had positive and significant impact on the knowledge level of farmers regarding citrus production and protection practices. T-test statistics showed highly significant (P<0.05) difference in knowledge level of FFS farmers and non-FFS farmers.

The biological functions of the BC047440 gene highly expressed by hepatocellular carcinoma (HCC) are unknown. The objective of this study was to reconstruct antisense eukaryotic expression vectors of the gene for inhibiting HepG2 cell proliferation and suppressing their xenograft tumorigenicity. The full-length BC047440 cDNA was cloned from human primary HCC by RT-PCR. BC047440 gene fragments were ligated with pMD18-T simple vectors and subsequent pcDNA3.1(+) plasmids to construct the recombinant antisense eukaryotic vector pcDNA3.1(+)BC047440AS. The endogenous BC047440 mRNA abundance in target gene-transfected, vector-transfected and naive HepG2 cells was semiquantitatively analyzed by RT-PCR and cell proliferation was measured by the MTT assay. Cell cycle distribution and apoptosis were profiled by flow cytometry. The in vivo xenograft experiment was performed on nude mice to examine the effects of antisense vector on tumorigenicity. BC047440 cDNA fragments were reversely inserted into pcDNA3.1(+) plasmids. The antisense vector significantly reduced the endogenous BC047440 mRNA abundance by 41% in HepG2 cells and inhibited their proliferation in vitro (P < 0.01). More cells were arrested by the antisense vector at the G1 phase in an apoptosis-independent manner (P = 0.014). Additionally, transfection with pcDNA3.1(+)BC047440AS significantly reduced the xenograft tumorigenicity in nude mice. As a novel cell cycle regulator associated with HCC, the BC047440 gene was involved in cell proliferation in vitro and xenograft tumorigenicity in vivo through apoptosis-independent mechanisms.

A resposta imune induzida por uma proteína recombinante de Leishmania (Leishmania) amazonensis de 33 kD (Larp33) foi avaliada em linfócitos de indivíduos vacinados com a Leishvacin® e em camundongos através de vacinação. Larp33 foi expressa em Escherichia coli após clonagem de um fragmento genômico de L. (L.) amazonensis de 2,2 kb no vetor pDS56-6His. Larp33 foi reconhecida por anticorpos IgG presentes no soro de indivíduos vacinados com Leishvacin® e induziu proliferação em linfócitos desses indivíduos em níveis comparáveis ao antígeno total de Leishmania. A análise por imunoblot indicou que Larp33 corresponde a uma proteína de aproximadamente 40 kD expressa em promastigotas de L. (L.) amazonensis e L. (Viannia) braziliensis. Hibridização com sonda de DNA correspondente a parte do fragmento clonado, também demonstrou que o gene codificador desta proteína é expresso em promastigotas destas duas espécies. Larp33 em associação com BCG (Bacille Calmette-Guerin) foi capaz de induzir 75% de proteção em camundongos C75BL/10 e BALB/c, suceptíveis à infecção por L. (L.) amazonensis. Linfócitos dos camundongos protegidos produziram IL-2 e IFN-<FONT FACE="Symbol">g</font> em resposta a Larp33. Nossos resultados indicam que Larp33 é imunogênica para linfócitos de indivíduos vacinados com Leishvacin ® e protetora para camundongos contra a infecção por L. (L.) amazonensis

In the search for Leishmania recombinant antigens that can be used as a vaccine against American Cutaneous Leishmaniasis, we identified a Leishmania (Leishmania) amazonensis recombinant protein of 33 kD (Larp33) which is recognized by antibodies and peripheral blood leukocytes (PBL) from subjects vaccinated with Leishvacin ®, Larp33 was expressed in Escherichia coli after cloning of a 2,2 kb Sau3A digested genomic fragment of L. (L.) amazonensis into the pDS56-6 His vector. Immunoblotting analysis indicated that Larp33 corresponds to an approximately 40-kD native protein expressed in promastigotes of L.(L.) amazonensis and L. (Viannia) braziliensis. Northern blots of total RNA also demonstrated that the gene coding for this protein is expressed in promastigotes of the major lineages of Leishmania causing American Cutaneous Leishmaniasis. Larp33 induced partial protection in susceptible mouse strains (BALB/c and C57BL/10) against L. (L.) amazonensis after vaccination using Bacille Calmette-Guerin (BCG) as adjuvant. In vitro stimulation of splenocytes from BALB/c protected mice with Larp33 elicited the secretion of IL-2 and IFN-<FONT FACE="Symbol">g</font>, suggesting that a Th1 cell-mediated protective response is associated with the resistance observed in these mice. As revealed by its immunogenic and antigenic properties, this novel recombinant antigen is a suitable candidate to compose a vaccine against cutaneous leishmaniasis