A novel mutation in the ABCD1 gene causes adrenomyeloneuropathy by impairing autophagy level / 中华神经科杂志

Objective:To report a Chinese family with a novel ABCD1 gene mutation at c.332T>G (p.V111G) site and discuss its clinical characteristics and molecular mechanism.Methods:The clinical data, laboratory examination, and imaging examination results were analyzed to make the clinical diagnosis of a middle-aged onset patient from the First Affiliated Hospital of Zhengzhou University in May 2017. High-throughput sequencing was used to discover a novel ABCD1 gene mutation. Sanger sequencing was used to find out whether other family members contain the same ABCD1 gene mutation. The pathogenicity of this mutation was explored by protein structure prediction and pathogenicity analysis. Adrenoleukodystrophy protein-green fluorescent protein (ALDP-GFP) and ALDP-GFP (V111G) plasmids were constructed and human embryonic kidney 293 cells were transfected, then immunofluorescence and Western blotting were used to explore the molecular mechanism of this mutation (completed in Henan Provincial People′s Hospital).Results:The proband (a 39-year-old male) was diagnosed as adrenomyeloneuropathy, a subset of X-linked adrenoleukodystrophy, with a novel heterozygous missense mutation in the ABCD1 gene at c.332T>G (p.V111G) site, and his mother and two daughters were all carriers. Protein structure prediction and pathogenicity results suggested that this mutation is pathogenic. Overexpression of ALDP-GFP (V111G) in the human embryonic kidney 293 cells resulted in a significant decrease in the expression levels of ALDP and the abnormal localization from the peroxisomal membrane to the cytoplasm, accompanied by significant down-regulation of LC3-Ⅱ/LC3-Ⅰ and beclin-1.Conclusion:c.332T>G (p.V111G) is a novel pathogenic mutation in the ABCD1 gene, which causes adrenomyeloneuropathy by impairing autophagy.

Interaction network of SARS-CoV-2 with host receptome through spike protein

Host cellular receptors are key determinants of virus tropism and pathogenesis. Virus utilizes multiple receptors for attachment, entry, or specific host responses. However, other than ACE2, little is known about SARS-CoV-2 receptors. Furthermore, ACE2 cannot easily interpret the multi-organ tropisms of SARS-CoV-2 nor the clinical differences between SARS-CoV-2 and SARS-CoV. To identify host cell receptors involved in SARS-CoV-2 interactions, we performed genomic receptor profiling to screen almost all human membrane proteins, with SARS-CoV-2 capsid spike (S) protein as the target. Twelve receptors were identified, including ACE2. Most receptors bind at least two domains on S protein, the receptor-binding-domain (RBD) and the N-terminal-domain (NTD), suggesting both are critical for virus-host interaction. Ectopic expression of ASGR1 or KREMEN1 is sufficient to enable entry of SARS-CoV-2, but not SARS-CoV and MERS-CoV. Analyzing single-cell transcriptome profiles from COVID-19 patients revealed that virus susceptibility in airway epithelial ciliated and secretory cells and immune macrophages highly correlates with expression of ACE2, KREMEN1 and ASGR1 respectively, and ACE2/ASGR1/KREMEN1 (ASK) together displayed a much better correlation than any individual receptor. Based on modeling of systemic SARS-CoV-2 host interactions through S receptors, we revealed ASK correlation with SARS-CoV-2 multi-organ tropism and provided potential explanations for various COVID-19 symptoms. Our study identified a panel of SARS-CoV-2 receptors with diverse binding properties, biological functions, and clinical correlations or implications, including ASGR1 and KREMEN1 as the alternative entry receptors, providing insights into critical interactions of SARS-CoV-2 with host, as well as a useful resource and potential drug targets for COVID-19 investigation.

Systemic analysis of tissue cells potentially vulnerable to SARS-CoV-2 infection by the protein-proofed single-cell RNA profiling of ACE2, TMPRSS2 and Furin proteases

Single-cell RNA profiling of ACE2, the SARS-CoV-2 receptor, had proposed multiple tissue cells as the potential targets of SARS-CoV-2, the novel coronavirus causing the COVID-19 pandemic. However, most were not echoed by the patients clinical manifestations, largely due to the lack of protein expression information of ACE2 and co-factors. Here, we incorporated the protein information to analyse the expression of ACE2, together with TMPRSS2 and Furin, two proteases assisting SARS-CoV-2 infection, at single cell level in situ, which we called protein-proofed single-cell RNA (pscRNA) profiling. Systemic analysis across 36 tissues revealed a rank list of candidate cells potentially vulnerable to SARS-CoV-2. The top targets are lung AT2 cells and macrophages, then cardiomyocytes and adrenal gland stromal cells, followed by stromal cells in testis, ovary and thyroid. Whereas, the polarized kidney proximal tubule cells, liver cholangiocytes and intestinal enterocytes are less likely to be the primary SARS-CoV-2 targets as ACE2 localizes at the apical region of cells, where the viruses may not readily reach. Actually, the stomach may constitute a physical barrier against SARS-CoV-2 as the acidic environment in normal stomach (pH < 2.0) could completely inactivate SARS-CoV-2 pseudo-viruses. These findings are in concert with the clinical characteristics of prominent lung symptoms, frequent heart injury, and uncommon intestinal symptoms and acute kidney injury. Together, we provide a comprehensive view on the potential SARS-CoV-2 targets by pscRNA profiling, and propose that, in addition to acute respiratory distress syndrome, attentions should also be paid to the potential injuries in cardiovascular, endocrine and reproductive systems during the treatment of COVID-19 patients.

Clinical Application of Immunofixtion Electrophoresis, Serum Protein Electrophoresis and Immunoglobulins and Light Chain Quantitative Analysis in the Diagnosis of Multiple Myeloma / 现代检验医学杂志

Objective To study the effectiveness of the laboratory diagnosis of multiple myeloma(MM)patients with immun-ofixtion electrophoresis (IFE),protein electrophoresis (SPE)and immunoglobulins and light chain quantitative analysis. Methods Selected 192 MM patients and 30 healthy controls during June 2012 to December 2013 and analyzed the results of IFE,SPE and immunoglobulins,and light chain quantitative analysis in MM patients.Results M protein bands were seen in 120 cases (62.5%)by using SPE and M protein were positive in 174 cases (90.6%)among the 192 MM patients by using IFE.IFE showed that IgG were maximum type of the M protein (106 of 192,55.2%).There were IgG-λ type 66 cases (34.4%),IgA type 36 cases (18.8%)and free light chain type 24 cases (12.5%).Immunoglobulins of different immuno-phenotypes had higher than the nomal group with serum immunoglobulin and light chain quantitative analysis (P <0.05). The detection rate was 67.7% (130/192).Whateverκ-type M protein orλ-type M protein,the ratio ofκ/λwas significantly abnormal (P <0.05).The detection rate was 85.4% (164/192).Conclusion The better detection rate of immunological techniques such as immunofixtion electrophoresis and immunoglobulins quantitative analysis might provide valuable basis for the diagnosis and treatment of MM clinically and prevent misdiagnosis.

Protective Effect of DADLE on Pulmonary Function in Rats with Sepsis / 华中科技大学学报(医学版)

Objective To study the protective effects of DADLE on pulmonary function in rats with sepsis and the mechanism.Methods Seventy-two SD rats were randomly divided into sham operated group(SO),septic group(SEP),and DADLEtreated group (DADLE).Sepsis model was induced by cecal ligation and puncture(CLP).In SO group,the abdomen was opened without any other treatment.In DADLE-treated group,DADLE(0.5 mg/ml) was administerd at a dose of 5 ml/kg by intravenous injection after CLP.Rats were sacrificed at the 2nd,6th and 10th h after CLP.Arterial blood was collected for blood gas analysis.The wet-to-dry lung weight ratio was measured,and the levels of TNF-a and IL-6 in blood were detected.The levels of MPO,MDA and ATP in lung tissue were determined.The pathologic changes of the lungs were examined under a light microscope.Results As compared with SEP group,PaCO_2,and PaO_2 were significantly increased(P<0.05),while the wet-to-dry ratio and levels of serum TNF-a and IL-6 were significantly reduced(P<0.05),and levels of MPO and MDA were significantly reduced(P<0.05) and content of ATP in lung tissue was significantly increased (P<0.05)in DADLE group.More severe pathological changes were found in SEP group than in DADLE group.Conclusion DADLE has pulmonary protective effects in rats with sepsis.