Serum antibody levels in COVID-19 patients six months after hospital discharge / 上海预防医学

ObjectiveTo investigate the level of serum antibodies in COVID-19 patients six months after discharge， and to provide data to evaluate the duration of IgM， IgG and neutralizing antibody titers in the patients. MethodsEnzyme-linked immunosorbent assay （ELISA） was used to determine the antibody levels of IgM and IgG， and the new coronavirus live virus neutralization test was used to detect the neutralizing antibodies in the plasma of 181 recovered patients. ResultsThe IgG positive rate was 92.27% （167/181） in COVID-19 patients six months after discharge， while the lgM positive rate was 28.18% （51/181）. Six months after hospital discharge， 117 recovered patients （64.64%） were positive for IgG antibodies and negative for IgM antibodies， indicating that they had produced stable antibodies. This result suggested that they had been infected with the new coronavirus （SARS-CoV-2） and were in the recovery stage. The positive detection rate of neutralizing antibodies was as high as 91.71%. ConclusionSix months after infection with SARS-CoV-2， IgG antibodies produced in the patients continue to exist， and the neutralizing antibodies maintain a high and stable level. Results of this study have important guiding significance for future research on the durability of new coronavirus antibodies.

Serum antibody levels in COVID-19 patients six months after hospital discharge / 上海预防医学

ObjectiveTo investigate the level of serum antibodies in COVID-19 patients six months after discharge， and to provide data to evaluate the duration of IgM， IgG and neutralizing antibody titers in the patients. MethodsEnzyme-linked immunosorbent assay （ELISA） was used to determine the antibody levels of IgM and IgG， and the new coronavirus live virus neutralization test was used to detect the neutralizing antibodies in the plasma of 181 recovered patients. ResultsThe IgG positive rate was 92.27% （167/181） in COVID-19 patients six months after discharge， while the lgM positive rate was 28.18% （51/181）. Six months after hospital discharge， 117 recovered patients （64.64%） were positive for IgG antibodies and negative for IgM antibodies， indicating that they had produced stable antibodies. This result suggested that they had been infected with the new coronavirus （SARS-CoV-2） and were in the recovery stage. The positive detection rate of neutralizing antibodies was as high as 91.71%. ConclusionSix months after infection with SARS-CoV-2， IgG antibodies produced in the patients continue to exist， and the neutralizing antibodies maintain a high and stable level. Results of this study have important guiding significance for future research on the durability of new coronavirus antibodies.

Construction of a VEGFR1 and VEGFR2 Bi-targeting oligopeptide-based fusion protein A7/G6-LDP and its anti-angiogenic activity / 中国药学杂志

OBJECTIVE: To construct a VEGFR1 and VEGFR2 bi-targeting oligopeptide-based fusion protein A7/G6-LDP and investigate its anti-angiogenic activity and the mechanism of action. METHODS: PCR and overlap PCR were used to construct the fusion protein A7/G6-LDP expression vector that consists of the gene encoding G6, A7, LDP, and the linker peptide. The product was purified by affinity chromatography and analyzed by SDS-PAGE and HPLC. The binding activity to endothelial cells was examined by ELISA and immunocytochemical staining. The inhibition of HMEC-1 proliferation was determined with CCK-8 assays. The phosphorylation of AKT and c-Raf was detected by Western blotting. HMEC-1 migration was determined using a wound healing assay and tube formation was measured after incubation on Matrigel. RESULTS: The data of DNA sequence confirmed that the A7/G6-LDP fusion protein was correctly constructed. The fusion protein was recovered in high yield (up to 20 mg·L-1) and high purity after His-tag purification. A7/G6-LDP bound to HMEC-1 and HUVEC, respectively; in addition, it inhibited endothelial cell proliferation more effectively than LDP alone when used higher concentration. Moreover, A7/G6-LDP disrupted the formation of endothelial tube structures and inhibited endothelial cell migration. The mechanistic study showed that A7/G6-LDP decreased the phosphorylation of AKT in HMEC. CONCLUSION: The engineered VEGFR1 and VEGFR2 bi-targeting oligopeptide-based fusion protein A7/G6-LDP effectively inhibits anti-angiogenesis. It might serve as a drug delivery carrier in targeted cancer therapy.

Effect of safflor yellow B on vascular endothelial cells injury induced by angiotensin-II / 药学学报

This study is to investigate protective effect of safflor yellow B (SYB) against vascular endothelial cells (VECs) injury induced by angiotensin-II (Ang-II). VECs were cultured and divided into six groups: control group, Ang-II group, Ang-II + SYB (1 micromolL(-1)) group, Ang-II + SYB (10 micromolL(-1)) group, Ang-II + SYB (100 micromolL(-1)) group and Ang- II + verapamil (10 micromolL(-1)) group. Except control group, all of VECs in other groups were treated with Ang- II at the final concentration of 0.1 micromolL(-1). Mitochondria membrane potential (MMP) and free calcium concentration ([Ca2+]i) were measured by laser scanning confocal microscopy, and mitochondria complex IV activity was detected by BCA method. The levels of reactive oxygen species (ROS) in VECs were analyzed by fluorescence detector and apoptosis of VECs was observed by flow cytometer. Caspase 3 was determined by Western blotting method. Comparing with control group, Ang-II was able to increase [Ca2+]i and ROS level, decrease MMP level, inhibit complex IV activity and enhance caspase 3 activity in VECs, as a result, enhance apoptosis of VECs. But SYB could significantly reduce the result induced by Ang- II relying on different dosages (P < 0.05 or P < 0.01). SYB was able to eliminate the effect of Ang-II on VECs via regulating [Ca2+]i, mitochondrial structure and function and inhibiting apoptosis.

Analysis of etiology of four pandemic influenza A (H1N1) virus outbreaks in Shandong province, in 2009 / 中华预防医学杂志

<p><b>OBJECTIVE</b>To isolate and identify the influenza virus that caused four influenza-like-illness outbreaks in Jining city of Shandong Province in 2009 and analyze the genetic characteristics of hemagglutinin (HA) and neuraminidase (NA) gene, the variation of these genes were studied.</p><p><b>METHODS</b>34 nasopharyngeal swabs from fever patients of four influenza-like-illness outbreaks were collected and diagnosed by real time quantitative RT-PCR method. The positive samples were incubated and cultured for virus. HA and NA genes of isolated pandemic influenza A (H1N1) virus were sequenced, the homology analysis was done with DNAStar software and the genetic evolution and amino acid substitutions were performed with Mega 4.0 software. The sequences were compared with WHO recommended vaccine virus, native reference virus.</p><p><b>RESULTS</b>Seventeen of 34 nasopharyngeal swabs were positive, 11 pandemic influenza A (H1N1) viruses were isolated and HA and NA genes of 7 strains were sequenced. Phylogenetic analysis for hemagglutinin and neuraminidase gene of Shandong outbreak strains showed that there were 98.4% - 99.6% and 99.2% - 100.0% sequence identity. Compared with WHO-recommended vaccine strain, the reference virus in mainland China strain, eleven amino acids were changed for HA protein, including position 38, 40, 56, 90, 100, 145, 172, 173, 220, 303 and 338, and 38, 40, 303 of HA protein were located in the antigenic determination C cluster, 172, 173 in the D cluster, 56 in the E cluster, site 40 of HA protein were glycosylated. In NA protein, seven amino acids were changed, including position 80, 106, 241, 248, 351, 369 and 386, site 40 of NA protein were glycosylated. No mutations of 275 in NA protein were found.</p><p><b>CONCLUSION</b>The HA and NA genes of the epidemic strains showed high homology, some mutations in the HA and NA proteins were found, the antigenic site and glycosylation site of some strains were changed during the epidemic process.</p>

Cloning and expression of the extracellular domain of 4-1BBL / 生物工程学报

RT-PCR was used to clone DNA fragment of the extracellular domain of 4-1BBL from human THP-1 cells (human monocyte), and the expression vector pAYZ4-1BBL was constructed by cloning the extracellular domain of 4-1BBL into the expression vector pAYZ. The extracellular domain of 4-1BBL was expressed in E. coli 16C9 and purified by affinity chromatography. SDS-PAGE and Western blot analysis showed that the relativae molecular weight of soluble 4-1BBL is 22kD which was consistent with the theoretically predicted value. So far as we know, it is the first time that the soluble expression of 4-1BBL in E. coli was achieved 4-1BBL induced a significant release of IL-2 in stimulated Jurkat cells after 48h incubation, especially in the presence of tumor cell. At the same time the apoptosis level of Jurkat cell reduce more than 50%. In conclusion, 4-1BBL may be useful in cancer immunotherapy.