Early detection of SARS-CoV-2 in circulating immune cells in a mouse model

SARS-CoV-2 infects the respiratory tract, lung and then other organs. However, its pathogenesis remains largely unknown. We used RareScope Fluorescence Light Sheet Microscopy (FLSM) and fluorescent in situ hybridization of RNA (RNA-FISH) to detect SARS-CoV-2 RNA and dissemination kinetics in mouse blood circulation. By RNA-FISH, we found that SARS-CoV-2 RNA-positive leukocytes, including CD11c cells, appeared as early as one day after infection and continued through day 10 post infection. Our data suggest that SARS-CoV-2-permissive leukocytes contribute to systemic viral dissemination, and RNA-FISH combined with FLSM can be utilized as a sensitive tool for SARS-CoV-2 detection in blood specimens.

UBXN3B Restricts Viral Pathogenesis by Maintaining Hematopoietic Homeostasis

Hematopoiesis is finely regulated to enable timely production of the right numbers and types of mature immune cells to maintain tissue homeostasis. Dysregulated hematopoiesis may compromise antiviral immunity and/or exacerbate immunopathogenesis. Herein, we report an essential role of UBXN3B in maintenance of hematopoietic homeostasis and restriction of immunopathogenesis during respiratory viral infection. Ubxn3b deficient (Ubxn3b-/-) mice are highly vulnerable to SARS-CoV-2 and influenza A infection, characterized by more severe lung immunopathology, lower virus-specific IgG, significantly fewer B cells, but more myeloid cells than Ubxn3b+/+ littermates. This aberrant immune compartmentalization is recapitulated in uninfected Ubxn3b-/- mice. Mechanistically, UBXN3B controls precursor B-I (pre-BI) transition to pre-BII and subsequent proliferation in a cell-intrinsic manner, by maintaining BLNK protein stability and pre-BCR signaling. These results reveal an essential role of UBXN3B for the early stage of B cell development.

Differential roles of RIG-I-like receptors in SARS-CoV-2 infection

The retinoic acid-inducible gene I (RIG-I) and melanoma differentiation-associated protein 5 (MDA5) are the major viral RNA sensors that are essential for activation of antiviral immune responses. However, their roles in severe acute respiratory syndrome (SARS)-causing coronavirus (CoV) infection are largely unknown. Herein we investigate their functions in human epithelial cells, the primary and initial target of SARS-CoV-2, and the first line of host defense. A deficiency in MDA5 (MDA5-/-), RIG-I or mitochondrial antiviral signaling protein (MAVS) greatly enhanced viral replication. Expression of the type I/III interferons (IFN) was upregulated following infection in wild-type cells, while this upregulation was severely abolished in MDA5-/- and MAVS-/-, but not in RIG-I-/- cells. Of note, ACE2 expression was ~2.5 fold higher in RIG-I-/- than WT cells. These data demonstrate a dominant role of MDA5 in activating the type I/III IFN response to SARS-CoV-2, and an IFN-independent anti-SARS-CoV-2 role of RIG-I.

The relationship between the characteristics of type 3 secretion system and biofilm of Pseudomonas aeruginosa in diabetic foot wound with antibiotic resistance / 中华内分泌代谢杂志

Objective:To explore the characteristics of type 3 secretion system and biofilm of Pseudomonas aeruginosa in diabetic foot wound, and to analyze the relationship between these factors, as well as to the antibiotic sensitivity.Methods:Thirty-three strains of Pseudomonas aeruginosa were collected from the foot wounds of diabetic foot inpatients in Tianjin Medical University Chu Hsien-I Memorial Hospital from February 1, 2018 to December 31, 2018. Thirteen strains of Pseudomonas aeruginosa were collected from non-diabetic wounds. All strains were tested for antibiotic sensitivity. The virulence genes exoS or exoU of Pseudomonas aeruginosa and the ability of biofilm formation were tested. The characteristics of exoS or exoU and biofilm of Pseudomonas aeruginosa were analyzed. Patients′ clinical outcomes were also analyzed.Results:Pseudomonas aeruginosa with exoS gene was the major pathogen, 90.9% found in diabetic foot group and 84.6% in control group, with no significant difference( χ2=0.54, P=0.46). The drug-resistant strains of Pseudomonas aeruginosa with exoS accounted for 16.7% in diabetic foot group and 18.2% in control group, also with no significant difference( χ2=0.18, P=0.83). There were 5 strains of Pseudomonas aeruginosa carrying exoU, 3 strains in diabetic foot group, of which 1 was resistant, 2 in control group, no resistant strain. Pseudomonas aeruginosa increased the ability of biofilm formation in diabetic foot group, accounting for 57.6%, and for resistant strains, 83.3% of them increased the biofilm formation ability. Two kinds of Pseudomonas aeruginosa produced different biofilms, but they were effectiveless for carbapenem antibiotics. The times of debridement ( P<0.01), time of antibiotic use ( P<0.01) were more in biofilm wound, but the healing rate reached 75%-90%. Conclusion:Pseudomonas aeruginosa secreting ExoS is the main one in the diabetic foot wound. The ability of Pseudomonas aeruginosa to produce biofilm in DF wound is increased. Biofilm is one reason for its antibiotic resistance. Multiple debridement combined with sensitive antibiotics is an effective method to remove biofilm.

Broad-spectrum virucidal activity of bacterial secreted lipases against flaviviruses, SARS-CoV-2 and other enveloped viruses

Viruses are the major aetiological agents of acute and chronic severe human diseases that place a tremendous burden on global public health and economy; however, for most viruses, effective prophylactics and therapeutics are lacking, in particular, broad-spectrum antiviral agents. Herein, we identified 2 secreted bacterial lipases from a Chromobacterium bacterium, named Chromobacterium antiviral effector-1 (CbAE-1) and CbAE-2, with a broad-spectrum virucidal activity against dengue virus (DENV), Zika virus (ZIKV), severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), human immunodeficiency virus (HIV) and herpes simplex virus (HSV). The CbAEs potently blocked viral infection in the extracellular milieu through their lipase activity. Mechanistic studies showed that this lipase activity directly disrupted the viral envelope structure, thus inactivating infectivity. A mutation of CbAE-1 in its lipase motif fully abrogated the virucidal ability. Furthermore, CbAE-2 presented low toxicity in vivo and in vitro, highlighting its potential as a broad-spectrum antiviral drug.

Effects of salvia miltiorrhiza polyphenols for injection and collagen sponge in diabetic foot / 中国生化药物杂志

Objective To investigate the effect of polyphenol of salvia miltiorrhiza polyphenols combined with collagen sponge in the treatment of diabetic foot.Methods A total of 115 patients with type 2 diabetic foot were randomly divided into treatment group (55 cases) and control group (60 cases).Salvia miltiorrhiza polyphenols and collagen sponge treatment group were treated with conventional treatment plus type 2 diabetes treatment of salvia miltiorrhiza polyphenol combined with collagen sponge;4 weeks as a treatment course, observation of 8 weeks, ulcer and other adverse reactions were recorded and compared.Results After 4 weeks of treatment, 17 cases (30.91%) with 30% or more reduction in foot ulcer area in treatment group,33 cases (60.00%) after 6 weeks treatment and the number of cases with 30% or more reduction in foot ulcer area,49 cases (89.09%) after 8 weeks of treatment, and the number of cases and the proportion of cases with 30% or more decrease of ulcer area in the control group after 4, 6 and 8 weeks were 12(20.00%), 22 (36.67%) and 39 (65.00%).There was no statistical difference between the two groups for the healing time of foot ulcer at three time points.After 8 weeks of treatment, the cure rate was 38.18% in salvia miltiorrhiza polyphenol combined with collagen sponge group. The effective rate was 50.91% and 45.00% for the two groups, the difference was not statistically significant; while the total effective rate was also significantly higher(89.09% vs.65.00%,P<0.01).Conclusion Salvia miltiorrhiza polyphenol combined with collagen sponge has a good effect on diabetic foot treatment, especially it can improve the cure rate, which is superior to the routine therapy.

Glucagon-like peptide 1 receptor agonist protects high-glucose inducedβcells apoptosis via inhibition of NOX2-dependent ROS production / 天津医药

Objective To investigate the possible mechanisms of glucagon-like peptide 1 receptor agonists (GLP-1Ra) protection against hyperglycemic induced beta cell apoptosis through depression of NOX2-dependent ROS production. Methods The rat model of type 2 diabetes (T2DM) was established by injecting small doses of streptozotocin (STZ) fol?lowed by 8-week high fat diet. The experimental animals were divided into three groups:normal control (N) group, diabetes (T2DM) group and GLP-1Ra group [treated with liraglutide 200 μg/(kg · d)for 12 weeks]. The blood glucose levels were compared before and after modeling, before treatment and 12-week after treatment with GLP-1Ra. The level of glycosylated hemoglobin (HbA1c) was detected by high-pressure liquid chromatography. Automatic biochemical analyzer was used to de?tect levels of aspertate aminotransferase (AST), creatinine (CR) and urea nitrogen (BUN). The apoptotic rates of islets were determined by TUNEL method and cleaved caspase 3 was detected by immunohistochemistry. DCFH-DA fluorescent probe was used to detect reactive oxygen species (ROS) levels of islets. Levels of NADPH oxidase (NOX) catalytic subunit (NOX 2) in islets were measured by immunohistochemistry. Results At the end of the study, glycemic control (average blood glucose/week and HbA1c) and lipid situation were improved significantly in the GLP-1Ra group than those of N group (P0.05). After application Apocynin for inhibition, there were no significant differences between three groups (P>0.05). The level of NOX2 was significantly lower in GLP-1Ra group compared to that of T2DM group (P<0.05). Conclusion GLP-1Ra can inhibit apoptosis ofβcells in diabetes rat, and the depression of NOX2-dependent ROS may be one of the important underly?ing mechanisms.

Effect of high glucose and anoxia on Amot expression in vascular endothelial cells with regard to its function in promoting angiogenesis / 中华内分泌代谢杂志

Objective To observe the effects of high glucose and anoxia on Amot expression in vascular endothelial cells (VECs),and explore its role in angiogenesis.Methods VECs were incubated with different glucose concentrations for 48 h,and then cultured at normal oxygen concentration or anaerobic condition for 24 h.The protein expressions of p130-Amot and p80-Amot were detected by Western blot.After Amot expression was downregulated in VECs by siRNA,wound healing experiments and angiogenesis experiments were performed to test the effect of decreased Amot expression on angiogenesis.Results pl30-Amot protein expressions in low glucose (5.5mmol/L) plus normal oxygen group and low glucose plus anaerobic group were higher than those in high glucose (30mmol/L) plus normal oxygen group,high glucose plus anaerobic group,middle glucose (15 mmol/L) plus normal oxygen group,and middle glucose plus anaerobic group (all P＜0.01).Compared with low glucose plus anaerobic group,p130-Amot expression was higher in low glucose plus normal oxygen group (P ＜ 0.01).However,the expression of p80-Amot showed no statistically significant difference among different groups (P＞0.05).Compared to the normal VECs,the cells with decreased Amot expression by siRNA exhibited an attenuated cell migration in the wound healing experiments and a lesser tube formation in the angiogenesis experiments.Conclusions High glucose exerts a more significantly negtive effect on the Amot expression than anoxia in VECs.The downregulation of Amot expression inhibits migration and angiogenesis of VECs.

Clinical Features and Antibiotic Sensitivity of Gram-Negative Diabetic Foot Osteomyelitis / 天津医药

Objective To investigate the clinical features and antibiotic susceptibility of osteomyelitis infected by Gram-negative bacteria (G-) in patients suffered from diabetic foot ulcers (DFU). Methods The clinical data of 91 DFU pa-tients accompanied with osteomyelitis (DFO) were retrospective studied. These patients hospitalized in the Tianjin Metabolic Diseases Hospital were divided into two groups, Gram-negative bacteria (G-) group (n=44) and Gram-positive bacteria (G+) group (n=42), respectively. The clinical features were compared between two groups. Logistic regression analysis was used to determine the risk factors for Gram-negative bactreial infection. The Gram-negative antibiogram was summarized. Results A total of 112 pathogens were isolated from 91 patients. G-bacteria were the most frequent pathogens (48.2%), following by G+ bacteria (47.3%) and fungi (4.5%). Pseudomonas aeruginosa was the majority of the G-bacteria. Comparing the two groups, the rate of antibiotic use within the previous 6 months was significantly higher in G-group (75.0%) than that of G+group (52.4%, P<0.05). There were no significant differences in the other indicators between two groups. The Logistic re-gression analysis revealed that the history of antibiotic use was the independent risk factor of G-bacterial infections in DFO patients. Antibiotics susceptibilities reflected G- bacteria were more prevalent to resist to cephalosporins and quinolonem, but sensitive to imipenem, ceftazidine and cefperazone-sulbactam. Conclusion Gram negative bacteria were not only the main pathogens isolated from DFO patients, but also frequently resistant to several popular antibiotics in China. The proper bacteria culture and antibiotic sensitivity test are especially emphasized to patients with DFU.

Study on the safety and effectiveness of an exercise prescription for patients with diabetic foot ulcer and peripheral arterial disease / 中华内分泌代谢杂志

Objective To investigate the effect of exercise prescription on foot perfusion and ulcer healing in patients with diabetic foot ulcer (DFU) accompanied with peripheral arterial disease (PAD).Methods Sixty patients with DFU and PAD were divided into exercise group and control group.The patients in the exercise group took exercise therapy training,ensuring that they could follow the exercise prescription.The patients in the control group had no exercise requirements.After the treatment for 12 weeks,ankle brachial index (ABI),transcutaneous oxygen tension (TcPO2),skin temperature,body mass index (BMI),HbA1C,and the ulcer healing rate in the two groups were compared,the satisfaction and compliance in the exercise group were evaluated,and the adverse events of the treatment were recorded.Results After the treatment for 12 weeks,the ABI,TcPO2,and foot skin temperature in the two groups increased and HbA1C decreased(P＜0.05 or P＜0.01).TcPO2 increased 5.25 mm Hg(1 mm Hg=0.133kPa),skin temperature increased 0.45℃,and BMI decreased 0.69 kg/m2 in the exercise group,while TcPO2 increased 2.59 mm Hg,skin temperature increased 0.28℃,and BMI increased 0.02 kg/m2 in the control group,showing significant differences in the three index changes between two groups (P＜0.01).The healing rate in the exercise group was higher than that in the control group (53.6％ vs 25.9％,P ＜0.05).The patients'compliance and satisfaction to the exercise therapy were 90％ and 94％,respectively.The incidence of adverse events in the exercise group had no causal relationship with the exercise prescription.Conclusion The exercise prescription in the study can improve the foot perfusion,and promote ulcer healing.It has the advantage of high safety,patients'compliance and satisfaction.