Dissolved oxygen concentration regulates human hepatic organoid formation from pluripotent stem cells in a fully controlled bioreactor

Developing technologies for scalable production of human organoids has gained increased attention for “organoid medicine” and drug discovery. We developed a scalable and integrated differentiation process for generation of hepatic organoid from human pluripotent stem cells (hPSCs) in a fully controlled stirred tank bioreactor with 150 ml working volume by application of physiological oxygen concentrations in different liver tissue zones. We found that the 20–40% dissolved oxygen concentration [DO] (corresponded to 30–60 mmHg pO₂ within the liver tissue) significantly influences the process outcome via regulating the differentiation fate of hPSC aggregates by enhancing mesoderm induction. Regulation of the [DO] at 30% DO resulted in efficient generation of human fetal-like hepatic organoids that had a uniform size distribution and were comprised of red blood cells and functional hepatocytes, which exhibited improved liver-specific marker gene expressions, key liver metabolic functions, and, more important, higher inducible cytochrome P450 activity compared to the other trials. These hepatic organoids were successfully engrafted in an acute liver injury mouse model and produced albumin after implantation. These results demonstrated the significant impact of the dissolved oxygen concentration on hPSC hepatic differentiation fate and differentiation efficacy that should be considered ascritical translational aspect of established scalable liver organoid generation protocols for potential clinical and drug discovery applications.     

Modulatory effects of statins on the autophagy: A therapeutic perspective

Autophagy is considered as an important mechanism for maintaining homeostasis and responsible for the degradation of superfluous or potentially toxic components and organelles. Autophagy impairment is associated with a number of pathological conditions, such as aging, neurological disorders, cancer, and infection. Autophagy also plays a significant role in cancer chemotherapy. The multiple cancer drugs have been notably developed with the strategy of autophagy modulation. Statins, 3-hydroxy-3-methyl-glutaryl-CoA inhibitors, are known due to their efficacy in decreasing low-density lipoprotein and extensively used for the management of cardiovascular diseases. Statins have other therapeutic and biological activities, such as antioxidant, anti-inflammatory, antitumor, and neuroprotective known as pleiotropic effects. It seems that statins are capable of targeting various signaling pathways in the induction of their great pharmacological effects. At the present study, we demonstrate the therapeutic effects of statins mediated via autophagy regulation.     

Nematodes associated with date palm and their control measures

Date palm, Phoenix dactylifera L., is dioecious and can be artificially pollinated by man, and one-third of all the dates of the world are grown in Iraq. In Egypt, there are about 12?million date palm trees grown in 99,867?feddans (fed.?=?4200?m²). Productivity is 1352,954?million?tons with yield 111.7?kg/tree. Plant parasitic nematodes associated with date palm are Criconemoides spp., Helicotylenchus spp., Hemicriconemoides spp., Hemicycliophora spp., Hoplolaimus spp., Meloidogyne incognita, Meloidogyne arenaria, Meloidogyne javanica, Pratylenchus brachyurus, Pratylenchus jordanensis, Pratylenchus coffeae, Pratylenchus neglectus, Pratylenchus thornei, Trichodorus spp., Tylenchorhynchus goffarti, Tylenchorhynchus latus and Xiphinema spp.; Meloidogyne incognita-infected roots of susceptible cultivar favoured giant cell and galls formation. Date palm roots infected with Pratylenchus penetrans showed puncture of epidermal cells and disarrangement of cortical cells with large empty abnormal cavities. As control measures, it is advised to; 1 – plant immune or resistant cultivars against pathogenic nematodes, 2 – use oil cakes or poultry manure as organic amendments and a nematicide, carbofuran.These were tested and found effective in the control of Helicotylenchus multicinctus and P. penetrans, 3 – treat nematode-infested date palm seedlings with hot water at a suitable temperature for a given period before transplanting to open field, 4 – plant nematode -free date palm seedlings, 5 – soil solarisation and tillage before planting, 6 – weed control, 7 – intercrop with nematode-resistant horticultural crops and 8 – induce resistance in susceptible date palm cultivars against root knot nematode.     

Phylogeography sheds light on the central–marginal hypothesis in a Mediterranean narrow endemic plant

Background and Aims

Understanding the factors that shape variation in genetic diversity across the geographic ranges of species is an important challenge in the effort to conserve evolutionary processes sustaining biodiversity. The historical influences leading to a central–marginal organization of genetic diversity have been explored for species whose range is known to have expanded from refugia after glacial events. However, this question has rarely been addressed for Mediterranean endemic plants of azonal habitats such as rocky slopes or screes. In this context, this comprehensive study examined molecular and field data from Arenaria provincialis (Caryophyllaceae), a narrow endemic plant of south-eastern France.

Methods

Across the whole geographic range, an investigation was made of whether high levels of abundance and genetic diversity (estimated from amplified fragment length polymorphism markers) are centrally distributed, to evaluate the relevance of the central–marginal hypothesis. Phylogeographic patterns inferred from chloroplast DNA (cpDNA) were used, applying Bayesian methods to test the influence of past biogeographic events. Multivariate analysis combining phylogeographic and ecological data was used to reveal the historical and ecological distinctiveness of populations.

Key Results

Despite the narrow distribution of A. provincialis, a high level of nucleotide variation is found within cpDNA loci, supporting its persistence throughout the Pleistocene period. The area characterized by the highest genetic diversity is centrally located. Structured phylogeography and Bayesian factor analysis supported the hypothesis that the central area of the distribution was the source of both westward and eastward migrations, probably during arid periods of the Pleistocene, and more recently was a crossroads of backward migrations. By contrast, the two areas located today at the range limits are younger, have reduced genetic diversity and are marginal in the ecological gradients.

Conclusions

This study highlights a case of strong population distinctiveness within a narrow range. Phylogeography sheds light on the historical role of the areas centrally situated in the distribution. The current range size and abundance patterns are not sufficient to predict the organization of genetic diversity.     

Reaction of sunflower varieties to Sclerotinia sclerotiorum under various inoculation methods,time of field evaluation and phylogenic stages of host

Sunflower cultivation is affected seriously by Sclerotinia sclerotiorum (Lib.) de Bary in Iran, particularly north-western areas. Because of economic and environmental harms by chemical control, it is necessary to develop cultivars with adequate genetic resistance for reduction of yield losses. The purpose of this study was to find an effective method of inoculation with S. sclerotiorum under field evaluations. Three stem-inoculation techniques including: 1 – mycelium plug, 2 – oxalic acid solution (OAS) and 3 – infested wheat seeds with Sclerotinia mycelium were employed under field conditions. Four genotypes including Ghalami (local variety in market), Confeta, Allstar and Master were used in this study. The lesion length, lesion width and lesion as up and down leading on the stem from inoculation site were measured after 3, 7, 10 and 14 days of inoculation. The analysis of variance showed significant difference between all employed techniques and incubation days after inoculation. Mycelial plug (MP) inoculation technique produced significantly more developed lesions on the treated stems. In spite of this effect, Master variety demonstrated reasonable resistance reaction against the disease. The progress of disease in wounded treatments was also faster than the non-wounded ones. And, the shortest time to obtain significant differences between varieties was 10?days after inoculation. By comparison of results of lesion length at flowering and seed-filling stages, the more obvious effectiveness of the disease was observed at the second stage. Finally, there were negative correlations between mean temperature and mean lesion length in all three inoculation methods.     

Studies on the transmission of some Fusarium spp. to potato progeny tubers from mother plants and its biological control

Fusarium spp. attack potato roots causing root-rot, damping-off and wilt disease in Assuit Governorate. Forty-five Fusarium isolates were isolated from F. nygamai, F. acutatum, F. solani, F. proliferatum, F. subglutinans, and F. oxysporum. Isolates were tested for their pathogenic capability on Burn potato variety during growing season 2007/2008. Isolates infect potato plants causing either damping-off or wilt symptoms. Isolates varied in their virulence. Role of potato tuber seed in the transmission of the causal pathogen to daughter using Electrophoresis. Protein profiles of the tested isolates divided into four sub-clusters at similarity levels 93.79, 91.55 and 92.62% while isolate of Fusarium profile No. 11 formed separate sub-clusters at similarity level 69.79%. F. nygamai and F. solani were notable exception because profile No. 4 of F. nygamai from roots and profile No. 4 from sprouts were almost identical (similarity level 96.81%); similarity level between profile No. 8 from roots and profile no/8 from sprouts was 95.44%. Results prove that F. nygamai and F. solani are potato tuber seed-borne fungus. T. harzianum, T. viride, T. longibrachiatum, G. virens and E. nigrum or its filtrate inhibited the growth of F. nygamai, F. acutatum, F. solani, F. proliferatum, F. subglutinans and F. oxysporum. The formulation of T. harzianum, T. longibrachiatum and G. virens against tested pathogenic fungi reduce disease incidence under greenhouse conditions.     

Co-limitation of potato growth by Potato Cyst Nematode (Globodera rostochiensis) and Rhizoctonia solani

Globodera rostochiensis and Rhizoctonia solani are the most important growth limiting factors influencing potato production in Iran. The effects of inoculation with Potato Cyst Nematodes (PCN) (0, 50, 75 and 100 cysts/3.5?kg soil) and R. solani (with or without inoculation) on potato growth and development were investigated in cultivars Santé and Marfona. Inoculation with R. solani induced severe damage, especially when inoculation was accompanied with high density of PCN. The damage caused by R. solani tended to increase with an increase in PCN density, especially in Marfona. In Santé, number of stems or branches per plant significantly increased by inoculation with R. solani, while in Marfona it was significantly affected either by R. solani inoculation or PCN density. In Santé, number of stolons per plant was significantly increased by PCN, but not by R. solani. In Marfona, however, the number of stolons per plant was significantly affected either by R. solani inoculation or by presence of PCN, but not affected by PCN density. The general effect of R. solani or PCN inoculation treatments on shoot, below-ground and total dry weight of potato was significant, but strongly affected by cultivar. In general, our study supports the synergistic interaction between R. solani and PCN and its moderation by the use of a resistant cultivar such as Santé.     

Cytokine single nucleotide polymorphisms in patients’ with gallstone: dose TGF-β gene variants affect gallstone formation?

Gallstone is a common biliary disorder with several risk factors. Immune responses and inflammatory cytokines are important in this disease; as a result, some cytokines can be detected in bile fluid. In this research, cytokine gene polymorphisms were studied, and their effects on gallstone formation were evaluated. On 158 gallstone patients and 254 normal subjects, by PCR- RFLP method, IL-4-C590T polymorphism and by ARMS-PCR method, IFN-γ T+874A, TNF-α-A308G, IL-6 G-174C and TGF-β T+869C variants were studied. Pathologic evaluations were done on surgical specimens. There were no significant differences in distribution of evaluated polymorphisms between patient group and normal control group (P > 0.05), except TGF-β +869T allele (P = 0.04, OR = 1.23, 95 % CI = 1–1.79) which was higher in patients with gallstone. Although the pro-inflammatory cytokines such as TNF-α and IL-6 may promote gallstone formation, in this study no significant correlation between TNF-α and IL-6 polymorphisms and gallstone formation was seen. It is taught that TGF-β may affect gallbladder cells to promote gallstone formation and higher producer TGF-β +869T allele can be a risk factor of gallstone disease, so further studies would be more elucidative.     

Cytokine gene polymorphism and graft-versus-host disease: a survey in Iranian bone marrow transplanted patients

Graft versus host disease (GVHD) is a major complication of bone marrow transplantation (BMT). Numerous studies have shown the potential role of cytokine genotypes in the occurrence of GVHD. In this retrospective, case–control study we aimed to investigate the association between 13 cytokine genes and acute GVHD (aGVHD) after HLA-identical sibling BMT in 91 Iranian subjects. Negative association was found between aGVHD and donor IL-10/GCC haplotype or donor IL-4Ra-A allele in the population study. When compared within the leukemia subgroup, we observed positive association between recipient IL-1α ?889/C allele and aGVHD. Also there were negative association between recipient IL-10/CAA haplotype and donor IL-4Ra/A allele and development of aGVHD. Among the different genotypes only donor IL-4Ra and donor IL-12 showed significant association. We conclude that several cytokine polymorphisms are positively and negatively associated with aGVHD in Iranian HLA matched siblings, of which IL-4Ra and IL-12 may play important roles.     

Multi-drug resistance-1 gene polymorphisms in nephrotic syndrome: Impact on susceptibility and response to steroids

Background

Role of multidrug resistance-1 (MDR-1) gene polymorphisms has not been clarified in nephrotic syndrome (NS). Additionally, researchers studied several genetic polymorphisms to explain their influence on different patients' responses to steroid; however the data were inconsistent. Therefore, we aimed to investigate the association of MDR-1 gene polymorphisms [C1236T, G2677T/A, C3435T] and haplotypes with susceptibility to childhood nephrotic syndrome, and whether they influence steroid response.

Methods

We detected MDR-1 gene polymorphisms using polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) in 138 NS patients and 140 age and sex matched healthy children.

Results

The frequencies of MDR1 G2677T/A GT, GA, TT + AA genotypes or T allele, MDR1 C3435T TT genotype, and T allele genotype frequencies were significantly increased in NS group. While no significant differences were observed in distributions of C1236T genotypes or allele between NS patients and healthy children. Moreover, steroid non-responder NS patients had significantly higher frequencies of MDR1 G2677T/A GT, GA, and TT + AA genotypes than steroid responsive NS patients. We observed also that NS patients with age less than 6 years old had increased frequencies of MDR1 G2677T/A GT, GA, TT + AA genotypes or T allele MDR1 C3435T CT, TT genotypes and T allele. Interestingly the frequency of the TGC haplotype of MDR1 was lower in the initial steroid responders than in non-responders NS patients. On the contrary, there were no any association between the MDR1 haplotypes with NS susceptibility and they did not influence renal pathological findings.

Conclusion

Our data suggested that MDR1 C3435T or G2677T/A gene polymorphisms are risk factors of increased susceptibility, earlier onset of NS, and steroid resistance.