Elucidation of mechanisms of action of Wei-Sheng-Fang-Yi-Bao-Dan in the treatment of COVID-19 and depression using network pharmacology and molecular docking

Purpose: To investigate the mechanisms of action of Wei-Sheng-Fang-Yi-Bao-Dan (WSFYBD) in the treatment of COVID-19 and depression using network pharmacology and molecular docking. Methods: First, the bioactive components and target genes of WSFYBD were retrieved from TCMSP database. The relevant gene targets of depression and COVID-19 were obtained from databases. The core WSFYBD genes for treatment were separately obtained by determining gene intersection. Cytoscape 3.8.0 software was used to draw the visual interactive networks. STRING database was employed to construct protein-protein interaction networks, while Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional enrichment analyses were used to determine the function and pathway of target genes via a Bioconductor/R. Finally, AutoDockTools software was employed for molecular docking. Results: A total of 105 potential bio-active components and 35 target genes of WSFYBD for COVID-19 therapy were identified. Also, 1905 GO entries (p < 0.05) and 158 related signal pathways (p < 0.05) for COVID-19 were obtained. Similarly, 114 potential bio-active components of WSFYBD and 127 potential therapeutic targets of depression were identified. Moreover, 1948 GO entries (p < 0.05) and 177 related signal pathways for depression were retrieved (p < 0.05). Docking results showed the main bio-active components were closely bound to the core targets. Conclusion: The mechanisms for treating COVID-19 show that WSFYBD directly acts on SARS-CoV-2 virus to prevent it from entering the host cell, or inhibits virus replication. Secondly, WSFYBD ameliorates depression by acting on key targets that control over-activated cytokines. Therefore, WSFYBD has potentials for the management of COVID-19 and depression

Comprehensive succinylome analyses reveal that hyperthermia upregulates lysine succinylation of annexin A2 by downregulating sirtuin7 in human keratinocytes

Background and Objectives: Local hyperthermia at 44°C can clear multiple human papillomavirus (HPV)-infected skin lesions (warts) by targeting a single lesion, which is considered as a success of inducing antiviral immunity in the human body. However, approximately 30% of the patients had a lower response to this intervention. To identify novel molecular targets for anti-HPV immunity induction to improve local hyperthermia efficacy, we conducted a lysine succinylome assay in HaCaT cells (subjected to 44°C and 37°C water baths for 30 min). Methods: The succinylome analysis was conducted on HaCaT subjected to 44°C and 37°C water bath for 30 min using antibody affinity enrichment together with liquid chromatography-tandem mass spectrometry (LC-MS/MS). The results were validated by western blot (WB), immunoprecipitation (IP), and co-immunoprecipitation (Co-IP). Then, bioinformatic analysis including Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment, motif characterization, secondary structure, and protein–protein interaction (PPI) was performed. Results: A total of 119 proteins with 197 succinylated sites were upregulated in 44°C-treated HaCaT cells. GO annotation demonstrated that differential proteins were involved in the immune system process and viral transcription. Succinylation was significantly upregulated in annexin A2. We found that hyperthermia upregulated the succinylated level of global proteins in HaCaT cells by downregulating the desuccinylase sirtuin7 (SIRT7), which can interact with annexin A2. Conclusions: Taken together, these data indicated that succinylation of annexin A2 may serve as a new drug target, which could be intervened in combination with local hyperthermia for better treatment of cutaneous warts

Research on the Cultural Tracing of the Patriarchal Clan System of Traditional Buildings in the Eastern Zhejiang Province, China, Based on Space Syntax: The Case Study of Huzhai in Shaoxing

The patriarchal clan system is an important prerequisite for the formation and development of Chinese traditional culture. The spatial layout and space usage of traditional buildings are intimately related to patriarchal culture. Thus, analyzing the spatial layout and usage is an effective way to trace the culture of traditional buildings. In this study, a typical traditional building named Huzhai in eastern Zhejiang was examined as an example. The spatial layout characteristics of Huzhai and different space usage relations corresponding to different users under the influence of the patriarchal clan system were investigated through “all lines” analysis in space syntax. In this process, traditional ritual activities were considered crucial for tracing the culture of traditional buildings in the eastern Zhejiang province. The results demonstrate that spatial layout and the usage of traditional buildings in the eastern Zhejiang province have led to “class” distinctions under the influence of patriarchal culture. The sacrificial activities of families further emphasize the class distinctions of building space. The differences in building space usage among different classes reflect the unequal distribution of social resources in China’s traditional feudal society. These differences reflect the inequality of space mastership and control among different classes that are a result of the unequal distribution of social resources in China

Global Deletion of TSPO Does Not Affect the Viability and Gene Expression Profile.

Translocator Protein (18kDa, TSPO) is a mitochondrial outer membrane transmembrane protein. Its expression is elevated during inflammation and injury. However, the function of TSPO in vivo is still controversial. Here, we constructed a TSPO global knockout (KO) mouse with a Cre-LoxP system that abolished TSPO protein expression in all tissues and showed normal phenotypes in the physiological condition. The birth rates of TSPO heterozygote (Het) x Het or KO x KO breeding were consistent with Mendel's Law, suggesting a normal viability of TSPO KO mice at birth. RNA-seq analysis showed no significant difference in the gene expression profile of lung tissues from TSPO KO mice compared with wild type mice, including the genes associated with bronchial alveoli immune homeostasis. The alveolar macrophage population was not affected by TSPO deletion in the physiological condition. Our findings contradict the results of Papadopoulos, but confirmed Selvaraj's findings. This study confirms TSPO deficiency does not affect viability and bronchial alveolar immune homeostasis

Generation of TSPO KO mice.

<p>(A) Schematic of generation of TSPO Floxed mice and conditional KO mice by Cre-LoxP System. Conditional TSPO KO mice were generated by targeting exon 2 and exon 3 of the mouse genomic locus, a NEO selection cassette Floxed by Frt sites. (B) Schematic of crossing program for TSPO global knockout mice, Protamine-Cre is a transgenic mouse where Cre recombinase expression is driven by sperm specific protamine promotor. (C) Genotyping of TSPO Floxed mice and KO mice by a TSPO genotyping primer pair 1 & 3, and Flp primers. (D) Genotyping of TSPO Floxed mice and KO mice by three TSPO genotyping primers 1, 2 and 3, and Protamine-Cre primers.</p

TSPO deletion does not affect viability by Het x Het and KO x KO breeding.

<p>(A) Birth rate of offspring of TSPO Het x Het breeding. (B) One litter Pups born 4 days from TSPO KO x KO breeding. (C) One litter pups born 9 days from TSPO KO x KO breeding. (D) Birth rate of offspring in TSPO KO x KO breeding. Chi-Squared Test was used to determine significant difference compared with Mendel’s Law.</p

TSPO KO mice show normal alveolar macrophage population.

<p>(A) FACS analysis of BALF and alveolar macrophages was identified as F4/80+, CD206+. (B) The percentages of alveolar macrophages in BALF from WT and TSPO KO mice are shown as the mean ± S.E.M. from four animals in each group (n = 4). NS, not significant by Student’s <i>t</i>-test.</p

TSPO expression was abolished in global KO mice without pathological changes.

<p>TSPO expression in different tissues from WT and KO mice were detected by western blotting (A) and IHC (B); (C) H&E staining of different tissues from WT and TSPO KO mice. Scale Bars, 100μm.</p