Biallelic mutations in spermatogenesis and centriole-associated 1 like (SPATC1L) cause acephalic spermatozoa syndrome and male infertility / 亚洲男科学杂志(英文版)

Acephalic spermatozoa syndrome is a rare type of teratozoospermia that severely impairs the reproductive ability of male patients, and genetic defects have been recognized as the main cause of acephalic spermatozoa syndrome. Spermatogenesis and centriole-associated 1 like (SPATC1L) is indispensable for maintaining the integrity of sperm head-to-tail connections in mice, but its roles in human sperm and early embryonic development remain largely unknown. Herein, we conducted whole-exome sequencing (WES) of 22 infertile men with acephalic spermatozoa syndrome. An in silico analysis of the candidate variants was conducted, and WES data analysis was performed using another cohort consisting of 34 patients with acephalic spermatozoa syndrome and 25 control subjects with proven fertility. We identified biallelic mutations in SPATC1L (c.910C>T:p.Arg304Cys and c.994G>T:p.Glu332X) from a patient whose sperm displayed complete acephalia. Both SPATC1L variants are rare and deleterious. SPATC1L is mainly expressed at the head-tail junction of elongating spermatids. Plasmids containing pathogenic variants decreased the level of SPATC1L in vitro. Moreover, none of the patient's four attempts at intracytoplasmic sperm injection (ICSI) resulted in a transplantable embryo, which suggests that SPATC1L defects might affect early embryonic development. In conclusion, this study provides the first identification of SPATC1L as a novel gene for human acephalic spermatozoa syndrome. Furthermore, WES might be applied for patients with acephalic spermatozoa syndrome who exhibit reiterative ICSI failures.

Research progress of iron metabolism and its mechanism in the treatment of heart failure / 药学学报

The pathogenesis of heart failure is a complex progression and associated with abnormal regulation of many signaling pathways. As a cofactor of hemoglobin, myoglobin, oxidative respiratory chain, DNA synthase and other important proteins, iron plays an indispensable role in myocardial energy metabolism. Recently, a large number of studies have shown that heart failure is related to the disorder of iron metabolism. Both iron deficiency and iron overload can lead to the development of a variety of cardiomyopathy, and even progress to heart failure. Iron metabolism could be a key target for the diagnosis, prevention and treatment of heart failure. Here, we review the basic process of iron metabolism and its mechanism in heart failure, expecting to provide new clues and evidence for the treatment of heart failure.

Effects of salidroside on the secretion of inflammatory mediators induced by lipopolysaccharide in the co-culture of rat alveolar macrophages and type II alveolar epithelial cells / 生理学报

The aim of the present study was to investigate the effect of salidroside (Sal) on inflammatory activation induced by lipopolysaccharide (LPS) in the co-culture of rat alveolar macrophages (AM) NR 8383 and type II alveolar epithelial cells (AEC II) RLE-6TN. CCK-8 colorimetric method was used to detect cell proliferation percentage. The enzyme-linked immunosorbent assay (ELISA) was used to determine the content of tumor necrosis factor alpha (TNF-α), macrophage inflammatory protein-2 (MIP-2) and interleukin-10 (IL-10) in the supernatant. Western blot was used to examine the expression levels of phosphorylated AKT (p-AKT) and total AKT protein. The results showed that pretreatment of RLE-6TN cells or co-culture of RLE-6TN and NR 8383 cells with 32 and 128 µg/mL Sal for 1 h, followed by continuous culture for 24 h, significantly increased the cell proliferation (P < 0.05). Compared with control group, 32 and 128 µg/mL Sal pretreatment significantly increased the ratio of p-AKT/AKT in RLE-6TN cells (P < 0.05). Pretreatment of 32 µg/mL Sal not only inhibited the secretion of TNF-α and MIP-2 by NR 8383 cells induced by LPS (P < 0.05), but also enhanced the inhibitory effect of RLE-6TN and NR 8383 cells co-culture on the secretion of TNF-α and MIP-2 by NR 8383 cells induced by LPS (P < 0.05). In addition, 32 µg/mL Sal pretreatment promoted LPS-induced IL-10 secretion by NR 8383 cells (P < 0.05), and enhanced the promoting effect of co-culture of RLE-6TN and NR 8383 cells on the IL-10 secretion by LPS-induced NR 8383 cells (P < 0.05). In conclusion, Sal may directly inhibit LPS-induced inflammatory activation of AM (NR 8383), promote the proliferation of AEC II (RLE-6TN) through PI3K/AKT signaling pathway, and enhance the regulatory effect of AEC II on LPS-induced inflammatory activation of AM.

Protective effect of synthetic salidroside on acute lung injury in rats / 生理学报

To study the protective effect and mechanism of synthetic salidroside on acute lung injury (ALI) induced by lipopolysaccharide (LPS), male Sprague-Dawley (SD) rats were randomly divided into saline control group, 3 mg/kg LPS model group, different doses of salidroside groups (5, 20 and 80 mg/kg), and 5 mg/kg dexamethasone group. Intratracheal LPS instillation was used to establish the ALI model 0.5 h after intraperitoneal injection of salidroside or dexamethasone, and the rats were sacrificed 6 h later. Lung wet/dry weight ratio (W/D) was calculated. Lung tissue pathology and lung injury score (LIS) were observed and evaluated through hematoxylin and eosin (HE) staining. The centrifugal sediment of bronchoalveolar lavage fluid (BALF) was used to count the polymorphonuclear leukocyte (PMN) number by Wright's staining, and the centrifugal supernatant of BALF was used to determine the contents of protein and inflammatory factors (TNF-α, IL-1β and IL-6). The contents of myeloperoxidase (MPO) and malondialdehyde (MDA) in lung tissue were determined. Western blot was used to detect the expression levels of phosphorylated and total nuclear factor kappa B (NF-κB)/p65 protein in lung tissue. The results showed that, compared with LPS group, the intervention of synthetic salidroside alleviated the pathological damage in lung tissue, decreased the LIS and lung W/D ratio (P < 0.05), reduced the PMN number, the contents of protein and inflammatory factors in BALF (P < 0.05), reduced the contents of MPO and MDA in lung tissue (P < 0.05), and inhibited the expression of p-NF-κB in lung tissue (P < 0.05). The results suggest that synthetic salidroside has a protective effect on ALI induced by LPS, and its mechanism is related to inhibiting the phosphorylation of NF-κB and reducing the aggregation of PMN in the lung.

Detection and clinicopathologic significance of microsatellite alteration and p53 mutation of neuroendocrine cells in colorectal adenocarcinoma / 中华病理学杂志

<p><b>OBJECTIVE</b>To study the possible clonal origin of neuroendocrine cells in colorectal adenocarcinoma.</p><p><b>METHODS</b>Twenty-six microsatellite loci were screened using laser capture microdissection, DNA extraction and whole genome amplification. Microsatellite instability (MSI) and loss of heterozygosity (LOH) in adenocarcinoma cells and neuroendocrine cells amongst 30 cases of colorectal carcinoma with neuroendocrine differentiation were detected using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP)-silver staining. The mutation status of p53 was evaluated by PCR-sequencing. The clonal origin of neuroendocrine cells in colorectal adenocarcinoma was determined.</p><p><b>RESULTS</b>Amongst the 30 cases studied, the prevalence of MSI was 16.9% while that of LOH was 8.5%. The rate showed no statistically significant difference between adenocarcinoma cells and neuroendocrine cells. In 6 cases, the microsatellite alteration was entirely consistent. In 23 cases, the rate of microsatellite alteration consistency was greater than that of inconsistency. In 1 case, the consistency and inconsistency rates were identical. There was statistically significant difference between consistency and inconsistency of microsatellite alteration. The prevalence of p53 mutation was 16.7% which was the same for both adenocarcinoma cells and neuroendocrine cells.</p><p><b>CONCLUSIONS</b>Adenocarcinoma cells and neuroendocrine cells in colorectal adenocarcinoma with neuroendocrine differentiation have similar biologic changes. It is likely that they are of identical origin.</p>

Prokaryotic expression and identification of human papillomavirus type 16 E5 protein / 南方医科大学学报

<p><b>OBJECTIVE</b>To construct the prokaryotic and eukaryotic expression vectors pET32/E5 and pcDNA3.1/E5 for transformation into E. coli BL21 and NIH(3)T(3) cells respectively to observe the expression of human papillomavirus type 16 E5 protein (HPV16 E5).</p><p><b>METHODS</b>HPV16 E5 gene was amplified by PCR from clinical isolates of HPV 16 and inserted into the plasmid pET32a(+) followed by digestion with BamH I and Hind III. The recombinant plasmid pET32/E5 was transformed into E. coli JM109 and selected with ampicillin. The positive clones containing the recombinant plasmid pET32/E5 were verified by restriction endonucleases BamH I and Xho and sequence analysis. The expression of HPV16 E5-TRX fusion protein in E. coli BL21(ED3) was identified by SDS-PAGE and Western blotting. The digestion product of BamH I and Xho was purified and inserted into the eukaryotic expression vector pcDNA3.1(+) to construct pcDNA3.1/E5, which was identified by sequencing and transfected into NIH3T3 cells. The NIH(3)T(3) cells with stable expression of HPV16 E5 were selected by G418 and confirmed by RT-PCR.</p><p><b>RESULTS</b>The pET32/E5 and pcDNA3.1/E5 vectors were constructed successfully. E.coli BL21(DE3) transformed by the recombinant plasmid pET32/E5 expressed HPV16 E5-TRX fusion protein efficiently. In the presence of 1 mmol/L IPTG at 28 degrees C, HPV16 E5-TRX recombinant protein accounted for about 10% of the total bacterial proteins. NIH3T3 cells stably expressing HPV16 E5 were harvested by selection with 250 g/ml of G418. HPV16 E5 gene from pcDNA3.1/E5-transfected NIH(3)T(3) cells was amplified by RT-PCR, and sequence analysis demonstrated the acquisition of the full-length gene fragment.</p><p><b>CONCLUSIONS</b>The prokaryotic and eukaryotic vectors for the HPV16 E5 gene have been successfully constructed. The acquisition of E .coli and NIH(3)T(3) cells with stable expression of HPV16 E5 protein may facilitate subsequent research of the biological properties and the transformation mechanism of HPV16 E5 protein on specific cells.</p>