The high-copy-number plasmid pcDNA3.1+ is unstable within S almonella typhimurium. A novel plasmid pmcDNA3.1+ was constructed by removin g the promoter sequence of ampicillin resistance gene (bla gene) in plasmid pcDNA3.1+. In contrast to pcDNA3.1+, pmcDNA3.1+ was stable within Salmonel la typhimurium SL7207 in LB medium with or without ampicillin. Further experi ments showed the ?-lactamase activity of Salmonella typhimurium SL7207(pmc DNA3.1+) was apparently lowered than that of Salmonella typhimurium SL7207( pcDNA3.1+) and the high ampicillin concentration was maintained longer in LB me dium culturing Salmonella typhimurium SL7207(pmcDNA3.1+). When mice were a dministered with Salmonella typhimurium SL7207(pmcDNA3.1+) intraperitoneall y, more than 95% of Salmonella cells separated from the spleen still harbore d the plasmid pmcDNA3.1+ 7 days later; but 99% of Salmonella cells lost the plasmid pcDNA3.1+ at day 3 in mice innoculated with Salmonella typhimurium SL7207(pcDNA3.1+). By lowering the expression of bla gene, the rapid deco mposition of ampicillin in LB medium was avoided and the metabolic pressure was relieved for the host cells. This method offers a solution for the problem of t he instability of high-copy-number plasmid within Salmonella typhimurium.

OBJECTIVETo explore the efficacy differences in early intervention of different acupuncture and moxibustion methods on gastrocnemius fatigue in rats induced by electrical stimulation.

METHODSFifty male SD rats were randomly divided into a control group, a model group, a hand acupuncture group, an electroacupuncture group and a moxibustion group, 10 rats in each group. Electrical stimulation of the sciatic nerve was given in the control group and gastrocnemius fatigue test was induced by electrical stimulation of the sciatic nerve in the model group after anesthesia without other treatment, but just take 6 times interval stimulation in the control group. The hand acupuncture group, the electroacupuncture group and the moxibustion group were treated with the corresponding acupoints stimulation method respectively for 20 min before gastrocnemius fatigue test, and Dazhui (GV 14) and Zusanli (ST 36) were selected. Immediately after gastrocnemius fatigue test, three or four gastrocnemius tissues at the same site on the right side were quickly taken for making specimen for transmission electron microscope (TEM). The changes of skeletal muscle ultrastructure of myofibrils, mitochondria, sarcoplasmic reticulum, glycogen particles were observed under TEM.

RESULTS(1) Muscle fibers disorder, partial mitochondrial vacuolization and glycogen particles smaller were shown in the model group. (2) No abnormalities were shown in the hand acupuncture group and the moxibustion group with mitochondrial morphology and number, which better than that in the model group, and glycogen particles increased. (3) Abnormal changes in morphology were shown in the electroacupuncture group with part of the muscle fibers derangement, Z line malalignment and a few mitochondria vacuolization.

CONCLUSIONHand acupuncture, electroacupuncture and moxibustion have the different effects on ultrastructure of gastrocoemius in rats. Acupuncture and moxibustion have shown good effects on the prevention and treatment of exercise-induced skeletal muscle cell and organelle damage and delaying exercise-induced fatigue.

Acupuncture Points ; Acupuncture Therapy ; Animals ; Glycogen ; metabolism ; Male ; Mitochondria ; metabolism ; ultrastructure ; Moxibustion ; Muscle, Skeletal ; metabolism ; ultrastructure ; Rats ; Rats, Sprague-Dawley

During the process of electroacupuncture (EA) therapy, whether there being a corrosive effect in ac- upuncture needles was observed. Acupuncture needles were inserted into a rabbit's acupoint to perform a 12-hour electrical stimulation with three types of common EA waveform; additionally two needles were put in 0.9% sodium chloride solution with 12-hour direct current. Afterwards, environmental scanning electron microscope was applied to detect the surface physical characteristics of acupuncture needles. As a result, after a 12-hour continued electri- cal stimulation with three types of common EA waveform in the rabbit, there was no corrosive effect in acupunc- ture needles; but the direct current could cause severe corrosion in acupuncture needles. It is believed that there is no corrosion effect on acupuncture needles in current EA treatment, and some accidents reported in literature may be related to quality of EA device or improper manipulation during the treatment.
Acupuncture Points ; Animals ; Corrosion ; Electroacupuncture ; instrumentation ; Male ; Needles ; adverse effects ; Rabbits

AIM: To determine the association between subfoveal choroidal thickness before therapy and therapeutic activity in diabetic macular edema.
METHODS: The current study was a retrospective study, which included 32 patients ( 32 eyes ) diagnosed with diabetic retinopathy and macular edema. All the patients were firstly treated with intravitreal injections of ranibizumab. Main outcome measures were included the subfoveal choroidal thickness, central macular thickness and best-corrected visual acuity ( BCVA) at preoperation and postoperative visit at 3mo.
RESULTS: After 3 monthly intravitreal injections of ranibizumab, the BCVA was significantly higher than that before therapy and accompanied with significantly reduced thickness of subfoveal choroid and central fovea of macula. Spearman analysis was revealed that a greater baseline subfoveal choroidal thickness was associated with a better BCVA (rs=0. 544, P=0. 036).
CONCLUSION:In the therapy of intravitreal injections of ranibizumab on diabetic macular edema, there seems to be a better BCVA in the patients with a greater baseline subfoveal choroidal thickness. Therefore, baseline subfoveal choroidal thickness may be a useful predictor for the therapy of diabetic macular edema.

Abstract: Objective　To analyze the nucleic acid detection results of severe acute respiratory syndrome corona virus-2 (SARS-CoV-2) by droplet digital PCR (ddPCR) and compare with the detection results of real-time fluorescence quantitative RT-PCR (qRT-PCR), so as to evaluate the advantages and disadvantages of detection, and to provide data support for optimizing the nucleic acid detection scheme of SARS-CoV-2. Methods　According to the SARS-CoV-2 specific primer probe published by the China Center for Disease Control and Prevention, a ddPCR detection method for SARS-CoV-2 was designed. One sample was selected for sensitivity test after gradient dilution; six respiratory virus nucleic acid positive samples including seasonal H3N2 influenza virus and SARS-CoV-2 positive samples were selected for specificity test; five SARS-CoV-2 positive samples were selected for repeatability test; in addition, 30 positive and 20 negative SARS-CoV-2 samples were selected for multiple clinical samples testing, and the results were analyzed and compared with those of qRT-PCR. Results　The ddPCR method can specifically detect SARS-CoV-2, and directly obtain the original copy number of the sample target gene to achieve accurate quantification; the sensitivity test of gradient dilution positive samples showed that qRT-PCR detected target genes in part of the 10-5 dilution of samples, and no target genes were detected in 10-6 dilution, while ddPCR detected all target genes in both 10-5 and 10-6 dilution of samples. The detection limit of ddPCR was two orders of magnitude higher than that of qRT-PCR, and the sensitivity was higher than that of qRT-PCR; in the comparison of the repeatability test results of the two methods, the coefficient of variation of ddPCR was 1.266%-11.814%, lower than 1.729%-26.174% of qRT PCR, and the repeatability was higher than qRT-PCR; among 50 clinical samples, 30 positive samples of confirmed cases of Coronavirus Disease 2019 (COVID-19) were detected by both methods, SARS-CoV-2 was successfully detected by both methods, and 20 negative samples of COVID-19 were detected by both methods, and the results were negative, with a coincidence rate of 100.00% (50/50). Conclusion　The ddPCR method can accurately quantify SARS-CoV-2 with strong specificity, and its sensitivity and repeatability are higher than those of qRT-PCR, but it also has certain detection limitations and is more suitable for the detection of low load samples. In the actual detection, the two methods can be reasonably combined to improve the detection accuracy.

This study aims to explore the impact on patient's liver and kidney function by different dosage of Kudiezi injection. This study retrospectively analyzed 15 228 patients' records from 18 nationwide general hospital information system (HIS). All patients were treated with Kudiezi injection, 1 956 patients that were given doses of > 40 mL, which is above the recommended dose, acted as the observation group. Fifty-five patients receiving the recommended dose of < 40 mL were the control group. Data about alanine aminotransferase (ALT), aspartate aminotransferase (AST), creatinine (Cr) and blood urea nitrogen (BUN) were collected before and after using Kudiezi injection, changes after treatment were outcomes. Also recorded were: age, costs, length of hospitalization and the patients' condition on admission. Propensity score method was used to balance 71 confounding variables such as gender, age, mortality, and costs. There were no significant difference on the four indexes between the two groups. It is hard to conclude that the use of Kudiezi injection over the recommended dose could influence the four indexes of liver and kidney from this data analysis. More conclusive evidence should be collected by further prospective study.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; Alanine Transaminase ; metabolism ; Aspartate Aminotransferases ; metabolism ; Blood Urea Nitrogen ; Drugs, Chinese Herbal ; adverse effects ; therapeutic use ; Female ; Hospital Information Systems ; Humans ; Injections ; Kidney ; drug effects ; metabolism ; Liver ; drug effects ; metabolism ; Male ; Middle Aged ; Prospective Studies ; Retrospective Studies ; Young Adult

Objective To analyze the impact factors for early renal damage in type 2 diabetic patients by the classification tree model.Methods A total of 601 patients with type 2 diabetes were enrolled.According to glomerular filtration rates and urine albumin quantification,the patients were divided into type 2 diabetes group (418 cases) and early diabetic renal damage group (183 cases).The clinical data of the patients were recorded to analyze the main influential factors for the microalbuminuria of type 2 diabetic patients using the Exhaustive CHAID classification tree algorithm.Results Six important explanatory variables were screened out by the classification tree model from the 34 candidate variables related to early renal damage,including fibrinogen,history of hypertension,retinopathy,Cys C levels,SBP and peripheral neuropathy.Elevated fibrinogen was the main factor.Conclusion The classification tree model can analyze the major influential factors of early renal damage in type 2 diabetic patients effectively,and it can help develop the prevention and treatment methods.

Objective To analyze the prevalence,risk factors of kidney disease in type 2 diabetic patients with KDIGO classification of chronic kidney disease,also to study cardiovascular and cerebrovascular diseases and death in these patients,so as to investigate the significance of the KDIGO classification system.Methods One thousand six hundred and forty-five type 2 diabetic patients who were in hospitalization from June 2008 to December 2012 were grouped according to the KDIGO classification of chronic kidney disease and the incidence of vascular disease was analyzed based on the classification.Clinical features were compared between patients with or without kidney disease.The risk factors of kidney disease and the death of diabetic patients were also investigated.Results There were 915 male and 730 female,aged a median (57.86±12.54) years with (6.35±6.30) years duration of diabetes mellitus among the 1645 cases,and 37.2％ of patients had concomitant kidney disease.According to the classi fi cation of CKD,patients in CKD group 3a,group 3b and CKD group 4-5 accounted for 5.7％,3.5％ and 7.6％,while 33.4％ of patients had proteinuria,among which 19.5％ with microalbuminuria,13.5％ with macroalbuminuria.On complications,patients with hypertension accounted for 49.5％,hyperlipidemia 67.7％,diabetic retinopathy 27.4％,cardiovascular and cerebrovascular diseases 18.5％ (coronary artery disease 16.5％,cerebrovascular diseases 8.8％).Statistical difference was detected in the incidence of diabetic retinopathy,coronary artery disease and cerebrovascular diseases between CKD group 3a and 3b (P ＜ 0.05).The duration of diabetes,concomitant hypertention especially with elevated systolic blood pressure,diabetic retinopathy and hyperuricemia were the independent risk factors for type 2 diabetic patients with kidney disease.Age,Scr,complicating cardiovascular and cerebrovascular diseases and advanced CKD stage were the independent risk factors for the death of type 2 diabetic patients with kidney disease.Conclusion KDIGO classification of chronic kidney disease enables better staging of kidney diseases in diabetic patients for management and prognosis.Diabetic patients have a higher prevalence of renal diseases and cardiovascular and cerebrovascular events than the general population.Early control of factors such as blood pressure and serum uric acid can delay the progression of kidney disease,and the predictive role of diabetic retinopathy should be emphasized.

Animals ; Helminthiasis ; pathology ; Humans ; Moniliformis ; pathogenicity

Objective To investigate the effect of tea polyphenols (TP) on renal damage in rats model induced by D-galactose. Methods Rats were injected with D-galactose (150 mg/kg?d),ip for 8 w,to induce renal damage. From the 3rd week,TP (150,75,37.5 mg/kg?d),aminoguanidine (150 mg/kg) and vitamin E (150 mg/kg) were administered with D-galactose for 6 w. After treatment,fasting blood glucose and 2 h blood glucose in oral glucose tolerance test were measured. The levels of HbA1C and fructosamine in serum,the activity of aldose reductase and content of advanced glycation end products (AGEs) in plasma and in kidney tissues and the activity of SOD,GSH-Px,and the contents of MDA in kidney tissues were measured,and 24h urinary protein,blood urea nitrogen (BUN) and serum creatinine (Cr) were detected. The apoptosis of renal cells were detected by flow cytometer. Results After treatment of D-galactose for 8 w,2h glucose level in oral glucose talerance test was increased significantly,the activity of aldose reductase and the content of AGES were increased significantly in blood. The levels of AGEs and MDA in renal tissues were also enhanced significantly. However,the activities of SOD and GSH-Px decreased. Additionally,the contents of 24h urine protein,BUN,Cr and the apoptotic rate of renal cells were increased significantly. High and middle dose of TP could can decrease the activity of aldose reductase in red blood cells,and inhibit the formation of glycation products in model rats induced by D-galactose. Also,TP could enhance the antioxidative activities and decrease the contents of AGEs and MDA in renal tissues. Mesnwhile,24h urine protein,BUN and Cr and the apoptotic rate of renal cells were increased significantly. Conclusion TP can inhibit glycation reaction induced by D-galactose and then protect renal from damage caused by glycation.