ABSTRACT
@#Abstract: Objective To clarify the long-term evolution of hepatitis B virus (HBV) quasi-species in HBsAg asymptomatic carriers in Long'an county, Guangxi. Methods ELISA was used to detect serological markers of HBV. Viral loads were measured by real time PCR. HBV DNA was extracted from serum by kits. The whole HBV genome was amplified using nested PCR and amplicons were sequenced by next-generation sequencing (NGS). These sequences from NGS were analyzed by the software like Mega. Results Serum samples were collected from 9 HBsAg asymptomatic carriers in Longan County，Guangxi at 4 different time points in 2004, 2007, 2013, 2019 or 2020. A total of 23 serum samples and 309 full-length gene quasi-species sequences were obtained, with an average amount of (0.18±0.07) G sequencing data for each sample. Genotype of 55.54%(5/9) the studied subjects underwent genotype conversion during the long-term evolution process of HBV quasi-species, and the genotyping results of the phylogenetic tree in the PreS/S region are in perfect agreement with the results of the whole genome analysis; recombinant B/C, I/C were found; the Sn ranged from 0 to 0.37 and the genetic diversity ranged from 0 to 0.11, respectively. A total of 21 special single nucleotide/amino acid mutations (7 in the S region, 2 in the X region, 3 in the PreC region and 9 in the BCP region) and 6 deletion mutations were detected, multiple mutations were found and no drug resistant mutations were found; 77.8%(7/9) of the HBV strains carried by the subjects in 2004 had double mutations at nt1 762(A→T) and 1 764(G→A) and a stop mutation at nt1 896(G→A); HBV mutations can be restored from the mutant type to the wild type and (or) vice versa without antiviral drug pressure, and The evolution rate of HBV genome was 2.03×10-5~3.50×10-3.Conclusion HBV genotype, recombinants, genetic complexity and diversity of HBV quasi-species can change over time during in natural infection. The transformation between HBV mutation type and wild type reduces the value of predicting clinical outcomes by genetic types and related mutations to some extent. The HBV genome evolution rate of asymptomatic carriers of HBsAg in Long'an County is very high.
ABSTRACT
Context: In cases of damaged corneal endothelium cells (CECs) of the eye, transplantation of cultured vascular endothelial cells (VECs) may be a viable method to restore transparency. Aims: To evaluate the viability of replacing damaged primate CECs with cultured allogeneic VECs. Subjects and Methods: Rhesus monkey VECs (RMVECs) were cultured and proliferating cells were labeled with bromodeoxyuridine (BrdU) in vitro. RMs of the experimental group (n = 6) underwent manual Descemettt membrane stripping with transplantation of RMVECs labeled with BrdU; those in the control group received manual Descemetnt membrane stripping without transplantation. Postoperative evaluations included the transparency and appearance of the corneal graft; distribution and ultrastructural changes of RMVECs on the inner surface of the cornea using scanning and transmission electron microscopy, and immunohistological identification of BrdU. Results: At 90 days postsurgery, the corneal grafts of the monkeys in the experimental group retained better transparency than those of the controls, without corneal neovascularization or bullous keratopathy. A layer of cells with positive BrdU staining was found on the posterior surface of the treated corneas in the experimental group, while there was no VEC structure in corneal grafts from the monkeys of the control group. Conclusions: RMVECs can grow on the posterior surface of the cornea without Descemet’s membrane. Cultured and transplanted RMVECs appeared similar in ultrastructure. VECs can provide a barrier to maintain corneal dehydration and transparency to some extent.