Baihe Dihuangtang Improves Hippocampal Neuron Damage in Anxious Depression Model Rats by Inhibiting NLRP3 Inflammasome Activation / 中国实验方剂学杂志

Objective:To investigate the anti-anxious depression mechanism of Baihe Dihuangtang from the NOD-like receptor thermal protein domain 3 （NLRP3） inflammasome. Method:Fifty SD rats were randomly divided into normal group， model group， venlafaxine group （13.5 mg·kg^-1）， Baihe Dihuangtang high and low dose group （16，4 g·kg^-1）， with 10 rats in each group. Chronic restraint stress for 28 days （6 h） combined with subcutaneous injection of corticosterone （30 mg·kg^-1） was used to establish induce an anxious depression model. From the 8th day of modeling， the rats in the normal group and the model group received distilled water， and those in groups with drug intervention were treated with corresponding drugs by gavage for 21 days. Elevated plus maze and open field test were used to evaluate the behavioral changes of rats. Enzyme- linked immunosorbent assay （ELISA） was used to detect serum and hippocampal interleukin-1<italic>β</italic> （IL-1<italic>β</italic>）， interleukin-6 （IL-6） and interleukin-18 （IL-18） levels. Western blot were used to detect the relative expression of hippocampal NLRP3， apoptosis-associated speck-like protein containing a CARD （ASC）， and Caspase-1. The pathological changes of the hippocampus were observed by hematoxylin-eosin（HE） staining， the average fluorescence intensity of NLRP3， ASC， and Caspase-1 was detected by immunofluorescence. The ultrastructure of neurons was observed under electron microscopy. Result:Compared with the normal group， the model group showed reduced total entries （TE）， the ratio of open-arm entries （OE%）， the ratio of open-arm times （OT%）， and the autonomous activity score （<italic>P</italic><0.01）， significant anxiety and depression-like behaviors， increased levels of IL-1<italic>β</italic>， IL-6， and IL-18 in the serum and hippocampus （<italic>P</italic><0.01）， elevated protein expression of NLRP3， ASC， and Caspase-1 （<italic>P</italic><0.01）， activated NLRP3 inflammasomes， and injured hippocampal neurons. Compared with the model group， the high-dose Baihe Dihuangtang group showed improved anxiety and depression-like behaviors （<italic>P</italic><0.01）， and decreased levels of IL-1<italic>β</italic>， IL-6， and IL-18 in the serum and hippocampus （<italic>P</italic><0.05，<italic>P</italic><0.01）， reduced protein expression of NLRP3， ASC， and Caspase-1 （<italic>P</italic><0.01）， and alleviated hippocampal neuron damage. Conclusion:Baihe Dihuangtang can improve neuronal damage in anxious depression by inhibiting the excessive activation of NLRP3 inflammasomes.

Chemistry research on natural product derivatives of dihydro-β-agarofuran sesquiterpenoids / 中国中药杂志

Dihydro-β-agarofuran sesquiterpenoids possess chemical diversity and biodiversity. A dihydro-β-agarofuran sesquiterpenoid with only hydroxyl groups has been prepared by basic hydrolysis of crude extract of Euonymus bungeanus with 0.4% yield. Twelve derivatives were available in esterification, oxidation, decarboxylation, etc. Extensive ~1H-NMR,~(13)C-NMR, MS spectroscopic analyses and single-crystal X-ray diffraction analysis with Cu Kα radiation indicated that eleven derivatives were new compounds. The results will provide reference for chemistry study on natural product derivatives of dihydro-β-agarofuran sesquiterpenoids.

Preparation of a polyclonal antibody against human LYZL4 and its expression in the testis / 中华男科学杂志

Objective@#To prepare a polyclonal antibody against human lysozyme-like protein 4 (LYZL4) expressed in the prokaryotic system and identify the distribution of LYZL4 in the testis.@*METHODS@#The full-length cDNA of LYZL4 was cloned into the pET32a plasmid and the expression of the recombinant LYZL4 (rLYZL4) was induced by IPTG. The rLYZL4 was purified by Ni-NTA and chitin affinity chromatography respectively and its bactericidal activity was observed by bilayer agar plate diffusion assay. The purified rLYZL4 was used as an immunogen to generate the polyclonal antibody, followed by examination of the antibody titer by ELISA and its specificity by Western blot. The distribution of LYZL4 in human tissue, sperm and seminal plasma was identified and its subcellular localization in the testis was determined by immunohistochemistry.@*RESULTS@#rLYZL4 was expressed efficiently in the prokaryotic system and exhibited no bacteriolytic activity against M. lysodeikticus and E. coli. The anti-rLYZL4 polyclonal antibody could bind the recombinant protein with a high sensitivity and specificity. LYZL4 was identified in the testis, epididymis and sperm protein extracts and localized in the acrosomal region of round and elongating spermatids.@*CONCLUSIONS@#An anti-rLYZL4 polyclonal antibody was successfully prepared using the prokaryotic expression system. LYZL4 was detected in the acrosomal region of round and elongating spermatids, suggesting an association with the structure and function of the acrosome.

Expression of human lysozyme-like protein 6 in the male reproductive system / 中华男科学杂志

<p><b>Objective</b>To study the expression of human lysozyme-like protein 6 (LYZL6) in the male reproductive system and its physiological role.</p><p><b>METHODS</b>The recombinant P. pastoris strain was cultured and induced with methanol to express LYZL6, followed by purification using chitin affinity chromatography. The bactericidal activity of the recombinant LYZL6 was observed by bilayer agar plate diffusion assay, and then the recombinant protein was used as an immunogen to generate polyclonal antibodies, whose specificity was examined by ELISA. The distribution of LYZL6 in the human tissue and semen was identified by Western blotting and the subcellular localization in the testis was investigated by immunohistochemistry.</p><p><b>RESULTS</b>At pH 5.6, recombinant LYZL6 exhibited a high bacteriolytic activity against M. lysodeikticus. ELISA analysis showed that the anti-LYZL6 polyclonal antibodies could bind the recombinant protein with a high specificity. Western blot manifested the expression of LYZL6 in the testis and epididymis, higher in the former than in the latter. LYZL6 was also detected in the sperm protein extract, while protein bands were not observed in the seminal plasma. Immunodetection with a specific antiserum localized the LYZL6 protein in the late spermatocytes and round spermatids.</p><p><b>CONCLUSIONS</b>LYZL6 has a higher bacteriolytic activity under low pH condition and is bound to spermatozoa after secreted in the testicular epithelia, suggesting that LYZL6 could act as a potential hydrolase for carbohydrates in zona pellucida penetration.</p>

Anti-tumor constituents from the roots of Actinidia valvata / 第二军医大学学报

Objective: To study the anti-tumor constituents from the roots of Actinidia valvata Dunn. Methods: Column chromatographic techniques were used to isolate and purify the chemical constituents of the plant and their structures were elucidated by spectroscopic analysis, including 1HNMR, 13CNMR and MS. The cytotoxic activities of some individual compounds were examined by MTT assay. Results: Ten triterpenoids: 2α, 3α, 24-trihydroxyurs-12-en-28-oic acid(1), asiatic acid(2), corosolic acid(3), 2α, 3α, 23, 24-tetrahydroxyurs-12-en-28-oic acid(4), 2α, 3α, 13β, 24-tetrahydroxyurs-11-en-28-oic acid-13-lactone(5), 2α, 3α, 24-trihydroxyolean-12-en-28-oic acid(6), 2α, 3α, 19α, 24-tetrahydroxyurs-12-en-28-oic acid(7), 2α, 3α, 24-trihydroxyurs-12, 20(30)-dien-28-oic acid(8), 2α, 3β, 24-trihydroxyurs-12-en-28-oic acid(9), and oleanolic acid (10), and one plant sterol β-citosterol(11) were isolated from the CHCl3 fraction of the roots of Actinidia valvata Dunn. The cytotoxic activities of five compounds (1, 2, 3, 4, and 7) against A549, LOVO and HepG2 cell lines were evaluated. Conclusion: Compounds 3-8 are isolated from this plant for the first time. Compounds 2 and 3 possess cytotoxic activity against LOVO and HepG2 cell lines, and the cytotoxic activity decreases with the increase of polarity of individual compound.