Humans ; Aged ; Lung Diseases ; Pneumonia/drug therapy* ; Adrenal Cortex Hormones/therapeutic use* ; Pulmonary Disease, Chronic Obstructive/drug therapy* ; Administration, Inhalation ; Community-Acquired Infections/drug therapy*

Objective To explore the role of Panax notoginseng saponins(PNS)in regulating the expression of sortilin and ATP-binding cassette transporter A1(ABCA1)in macrophages,and the effect of PNS on inhibiting formation of foam cells and the potential mechanism of PNS adjusting sortilin expression and cholesterol metabolism.Methods The macrophages were divided into five groups as follows:group A(only added with cell culture),group B(transfected with negative control lentivirus),group C(transfected with lentivirus-mediated sortilin overexpression),group D(transfected with lentivirus-mediated sortilin overexpression+60 μg·mL-1PNS),group E(transfected with lentivirus-mediated sortilin overexpression+10 μmol·L-1 mitogen-activated protein kinase kinase(MEK)inhibitor PD98059+60 μg·mL-1 PNS).The protein contents of sortilin,ABCA1,extracellular signal-regulated kinase(ERK)and phosphorylated-ERK(p-ERK)were evaluated with Western blot.All the cells in five groups were cultured with 50 μg·mL-1ox-LDL to form foam cells.The lipid in macrophages was investigated with red O assay.Results The relative expression levels of sortilin protein were 1.00±0.08,0.91±0.15,2.28±0.13,1.62±0.09 and 2.01±0.08;the relative expression levels of ABCA1 protein were 1.00±0.01,0.92±0.07,0.29±0.04,0.66±0.09 and 0.44±0.07;the ratios of p-ERK/ERK protein were 1.00±0.09,0.92±0.05,1.03±0.12,2.00±0.12 and 1.64±0.14;the contents of lipid in macrophages were(4.82±2.19)％,(6.70±0.88)％,(44.56±4.15)％,(27.66±3.25)％and(41.67±5.45)％.Except the ratios of p-ERK/ERK,the other parameters between group C and group A were statistically significant difference(all P＜0.01).Meanwhile,there were also statistically significant difference between group D and group C as well as group D and group E(P＜0.05,P＜0.01).Conclusion PNS inhibits the lipid accumulation in macrophages through upregulating ABCA1 and downregulating sortilin,and ERK signaling pathway may be as one of important mechanisms influencing the expression of sortilin and ABCA1 mediated by PNS.

Objective To study the effect of Hedysarum polysaccharides(HPS)on the expression of transforming growth factor-β,(TGF-β1),smad homologue 3 recombinant protein(smad3)and smad7 in renal tissue of db/db mice with diabetic nephropathy(DN).Methods According to their body weight,6-week-old male db/db mice were randomly divided into 5 groups:model group(0.9％NaCl 0.2 mL·d-1),positive control group(22.75 mg·kg-1·d-1 irbesartan)and experimental-H,-M,-L groups(200,100,50 mg kg-1·d-1 HPS),with 10 mice in each group;another 10 SPF grade male C57BL/6 mice of the same week were selected as normal group(0.9％NaCl 0.2mL·d-1).The mice in the 6 groups were given intragastric administration once a day for 12 weeks.The blood glucose concentration of mice was measured before treatment and at the 4th,8th and 12th week after treatment.The expression levels of TGF-β1,smad3 and smad7 were detected by Western blotting.Results After treatment,the blood glucose levels of the model group was significantly higher than those of the normal group(all P＜0.01);compared with the model group,the levels of blood glucose in the experimental-H,-M groups decreased significantly,and the differences were statistically significant(P＜0.05,P＜0.01).The relative expression levels of TGF-β,protein in normal group,model group,positive control group and experimental-H,-M groups were 0.71±0.16,1.66±0.18,1.00±0.17,0.88±0.15 and 1.23±0.15;the relative expression levels of smad3 protein were 0.89±0.32,2.26±0.35,1.24±0.31,1.05±0.30 and 1.67±0.35;the relative expression levels of smad7 protein were 1.66±0.03,0.60±0.03,1.10±0.07,1.48±0.08 and 0.97±0.09;there were statistically significant differences between the experimental-H,-M groups and the model group(P＜0.05,P＜0.01).Conclusion Hedysarum polysaccharides can improve renal fibrosis and delay the development of diabetic nephropathy by regulating the level of blood glucose,inhibiting TGF-β1,smad3 and increasing the expression of smad7.

Objective To investigate the genetic causes of auditory neuropathy with optic atrophy in a family.Methods The proband's medical history and family history were inquired in detail,and relevant clinical examina-tions were performed to confirm the diagnosis of auditory neuropathy with optic atrophy,and the genetic pedigree of the family was drawn.Peripheral blood of proband(Ⅲ-7)was collected for whole exome sequencing,and the patho-genicity of the detected mutations were interpreted.Blood samples of proband's wife(Ⅲ-8),eldest daughter(Ⅳ-7),second daughter(Ⅳ-9)and son(Ⅳ-10)were tested for mutation sites by Sanger sequencing.Combined with clinical manifestations and examination results,the family was studied.Results The genetic pattern of this family was autosomal dominant.The proband showed decreased visual acuity at the age of 19,bilateral sensorineural deaf-ness at the age of 30,and decreased speech recognition rate.Among 20 members of the family of 5 generations,10(2 deceased)showed similar symptoms of hearing and visual impairment.Proband(Ⅲ-7),eldest daughter(Ⅳ-7)and son(Ⅳ-10)underwent relevant examination.Pure tone audiometry showed bilateral sensorineural deafness.ABR showed no response bilaterally.The 40 Hz AERP showed no response in both ears.OAE showed responses in some or all of the frequencies.No stapedial reflex was detected.The eye movement of Ⅲ-7 and Ⅳ-10 were reasona-ble in all directions,and color vision was normal.Ocular papilla atrophy was observed in different degrees in fundus examination.OCT showed thinning of optic disc nerve fibers in both eyes,and visual evoked potential showed pro-longed P100 wave peak.They were diagnosed as hereditary auditory neuropathy with optic atrophy.A mutation of the OPA1 gene c.1334G＞A(p.Arg445His,NM_015560.2)at a pathogenic locus on chromosome 3 was detected by whole exon detection in Ⅲ-7.The results of generation sequencing analysis showed that the OPA1 gene c.1334G＞A(p.Arg445His,NM_015560.2)mutation of chromosome 3 was also found in Ⅳ-7 and Ⅳ-10.Meanwhile,the gen-otypes of Ⅲ-8 and Ⅳ-9 were wild homozygous,that is,no mutation occurred.Conclusion The OPA1 c.1334G＞A(p.Arg445His,NM_015560.2)mutation site might be the pathogenic mutation in this family.

Objective To prepare mouse monoclonal antibodies against the ectodomain of E2 (E2ecto) glycoprotein of Western equine encephalitis virus (WEEV). Methods A prokaryotic expression plasmid pET-28a-WEEV E2ecto was constructed and transformed into BL21 (DE3) competent cells. E2ecto protein was expressed by IPTG induction and presented mainly as inclusion bodies. Then the purified E2ecto protein was prepared by denaturation, renaturation and ultrafiltration. BALB/c mice were immunized with the formulated E2ecto protein using QuickAntibody-Mouse5W as an adjuvant via intramuscular route, boosted once at an interval of 21 days. At 35 days post-immunization, mice with antibody titer above 1×10⁴ were inoculated with E2ecto intraperitoneally, and spleen cells were fused with SP2/0 cells three days later. Hybridoma cells secreting specific monoclonal antibodies were screened by the limited dilution method, and ascites were prepared after intraperitoneal inoculation of hybridoma cells. The subtypes and titers of the antibodies in ascites were assayed by ELISA. The biological activity of the mAb was identified by immunofluorescence assay(IFA) on BHK-21 cells which were transfected with eukaryotic expression plasmid pCAGGS-WEEV-CE3E2E1. The specificity of the antibodies were evaluated with E2ecto proteins from EEEV and VEEV. Results Purified WEEV E2ecto protein was successfully expressed and obtained. Four monoclonal antibodies, 3G6G10, 3D7G2, 3B9E8 and 3D5B7, were prepared, and their subtypes were IgG2c(κ), IgM(κ), IgM(κ) and IgG1(κ), respectively. The titers of ascites antibodies 3G6G10, 3B9E8 and 3D7G2 were 10⁵, and 3D5B7 reached 10⁷. None of the four antibody strains cross-reacted with other encephalitis alphavirus such as VEEV and EEEV. Conclusion Four strains of mouse mAb specifically binding WEEV E2ecto are successfully prepared.
Horses ; Animals ; Mice ; Encephalitis Virus, Western Equine ; Ascites ; Immunosuppressive Agents ; Antibodies, Monoclonal ; Immunoglobulin M

Lipopolysaccharides (LPS) are major outer membrane components of Gram-negative bacteria. Unlike most Gram-negative bacteria, Acinetobacter baumannii can still survive and acquire polymyxin resistance after complete loss of LPS. Previous studies of LPS-deficient Acinetobacter baumannii mostly focused on LPS-deficient strains induced by polymyxin, whose background was complex and unstable. To investigate LPS loss-mediated polymyxin resistant-Acinetobacter baumannii, this study constructed a stable and clear background LPS-deficient strain by knocking out lpxC gene in Acinetobacter baumannii ATCC 19606 with CIRSPR/Cas9, and then studied the phenotypic changes of the lpxC-deficient strain including morphology, growth rate, antibiotic susceptibility, virulence, membrane permeability, and membrane potential. Animal experiments were approved by the Animal Care and Welfare Committee Institute of Medicinal Biotechnology, CAMS and PUMC (approval number: IMB-20240119D₉). The results indicated that the lpxC gene of Acinetobacter baumannii ATCC 19606 was successfully knocked out. After losing the lpxC, the strain underwent the morphological change from rod-shaped to spherical. Furthermore, it leads to reduced growth rate, enhanced membrane permeability, decreased membrane potential, lower virulence, and increased antibiotic susceptibility to β-lactams, quinolones, aminoglycosides, macrolides, glycopeptides. The lpxC deletion results in significant changes in membrane homeostasis and adaptability of Acinetobacter baumannii. Understanding the phenotypic changes of colistin-resistant Acinetobacter baumannii mediated by LPS loss is useful for exploring the resistance mechanism of Acinetobacter baumannii and developing new therapeutic strategies.

Objective To investigate the detection rate,epidemiology and resistance mechanism of methicil-lin-resistant Staphylococcus aureus(MRSA)in a hospital in recent 5 years.Methods A total of 762 strains of non repetitive Staphylococcus aureus detected from 2016 to 2020 in a hospital were collected retrospectively.Methicillin-sensitive Staphylococcus aureus(MSSA)was 392 strains(MSSA group)and 370 strains caused by MRSA(MRSA group),and 95 strains of MRSA isolated in 2020 were further used for resistance mechanism.Staphylococcus aureus was identified and tested for drug sensitivity by Vitek 2 automatic microbial system.Molecular epidemiological typing was determined by multilocus sequence typing.The biofilm formation was performed by crystal violet staining.PCR amplification was used to detect drug resistance genes,virulence genes and biofilm related genes,and logistic regression analysis was used to investigate the independent risk factors of its occurrence.Results The detection rate of MRSA in past five years was 48.56％,mainly was from pus samples and secretion samples(38.38％,33.51％respectively).MRSA was found in the general sur-gery(18.65％)and otorhinolaryngology(12.70％).ST88 was the most common multilocus sequence typing(37.89％),and followed by ST951(24.21％).Moderate biofilm formation was the most common,accounting for 74.73％.Multivariate regression analysis showed that compared with MSSA group,hypoproteinemia,en-docrine system diseases,wound infection and history of antibiotic use within six months were the independent risk factors for infection in MRSA group.Compared with the control group,hospital transfer,wound infection and tumor were independent risk factors for infection in MRSA group(P＜0.05).Conclusion The detection rate of MRSA in a hospital is high,and the carrying rate of various drug-resistant genes is high.The hospital should pay attention to the prevalence of MRSA and related risk factors,so as to prevent it early.

Objective To establish a mouse model of type Ⅰ tricho-hepato-enteric syndrome(THES)induced by Ttc37 deficiency.Methods Ttc37 flox strain was established by site-specifically inserted loxP sites into Ttc37 gene via CRISPR/CAS9 technology.Ubiquitously expressed CAG-Cre was introduced for all-tissue removal of Ttc37 in Ttc37flox/flox;CAG-Cre mice.The knock-out effect was confirmed by fluorescence quantitative PCR and Western blot.Phenotypic evaluations were conducted in 8-week-old mice including hematoxylin-eosin staining of skin,spleen,liver,bladder,and gastrointestinal tract(GI),serum enzyme activity assay of aspartate aminotransferase(AST)and alanine aminotransferase(ALT),measurement of serum hemoglobin level,and ELISA for IgG and IgM level upon antigen immunization.Results Similar to type Ⅰ THES patients,Ttc37flox/flox;CAG-Cre mice exhibited impaired development of hair shaft,epidermis,B cell and eyes,while liver,GI,bladder and serum hemoglobin level seemed normal under unstressed condition.Conclusion A novel mouse model of typeⅠ THES was constructed successfully,which was applicable for pathological study.

Objective To investigate the effect and safety of common electric knife clamp coagulation technique in rabbit thyroidectomy.Methods According to the random number table method,12 New Zealand rabbits were divided into the clamp coagulation group and the ultrasonic scalpel group,with 6 rabbits in each group.The middle part of the thyroid gland in the clamp coagulation group was severed by common electric knife clamp coagulation,while that in the ultrasonic scalpel group was severed by ultrasonic scalpel electrocoagulation.The postoperative conditions of rabbits in the two groups were observ.The severed thyroid tissue was stained with hematoxylin-eosin(HE),and its histopathology after thermal damage was observed under the light microscope.The scope of thermal damage was determined.On the 1st,3rd and 7th day after operation,the auricular venous blood of all rabbits was collected to assess the serum interleukin-6(IL-6)and C-reactive protein(CRP)levels by enzyme-linked immunosorbent assay.The rabbits were killed on the 7th day after operation,and the residual thyroid glands were removed and stained by HE.The pathological changes and inflammatory cell infiltration were observed under the light microscope.Results The rabbits in the 2 groups survived well after operation,and the operative area healed well.No obvious effusion,blood clot,bleeding,incision infection or other complications were found in the residual cavity.Under the light microscope,the surface of the incisal margin of the thyroid gland showed obvious lesions.In the injured area,some cell structures were damaged with coagulated necrosis,some follicles were ruptured,and the contents inside were solidly concentrated and deeply stained.The cytoplasmic eosinophils in parafollicular cells increased,and nuclear pyknosis,fragmentation,and even dissolution occurred.The thermal damage ranges of thyroid tissues in the ultrasonic scalpel and clamp coagulation groups were(0.72± 0.10)mm and(0.88±0.11)mm,respectively.The range of thermal damage in the clamp coagulation group was significantly greater than that in the ultrasonic scalpel group(t=-2.740,P＜0.05).On the 1st,3rd and 7th day after surgery,there was no significant difference in the levels of serum CRP and IL-6 between the two groups(P＞0.05).The serum IL-6 levels in both groups on the 3rd and 7th day after surgery were significantly higher than those on the 1st day after surgery(P＜0.05).There was no significant difference in serum IL-6 level on the 3rd and 7rd day after surgery in the two groups(P＞0.05).Thyroid follicular atrophy,glia reduction,follicular epithelial hyperplasia,collagenization and hyperplasia of interstitial fibers were observed in the residual thyroid sections of both groups.No obvious inflammatory cell infiltration was observed.Conclusion In rabbit thyroidectomy,it is safe to remove the thyroid gland using the common electric knife clamp coagulation technique.In terms of preventing thermal damage,the ultrasonic scalpel is better than the common electric knife clamp coagulation technique,but the thermal damage to thyroid tissues caused by the common electric knife clamp coagulation technique is within the safe operating range.

Objective:To analyze the external quality control assessment results of salt iodine, urine iodine, and water iodine in iodine deficiency disorders (IDD) laboratories at all levels in Shandong Province, and provide reliable quality assurance for monitoring and prevention of IDD.Methods:The external quality control assessment of salt iodine, urine iodine and water iodine in provincial, municipal and county levels IDD laboratories in Shandong Province from 2017 to 2021 were conducted, and feedback rate and qualification rate were calculated.Results:From 2017 to 2021, the feedback rates and qualification rates of external quality control assessment of salt iodine, urine iodine and water iodine at provincial and municipal levels IDD laboratories in Shandong Province were all 100.0%. The feedback rates of county level laboratories participating in the salt iodine external quality control assessment were all 100.0%. The qualification rate in 2021 was 99.3% (135/136), and the other four years were all 100.0%. The feedback rates of county level laboratories participating in the urine iodine external quality control assessment were all 100.0%. The qualification rates in 2017 and 2021 were 98.4% (122/124) and 97.1% (132/136), respectively, and the other three years were all 100.0%. In 2021, the county level laboratories in Shandong Province participated in the water iodine external quality control assessment for the first year, and the feedback rate and qualification rate of 69 laboratories were both 100.0%.Conclusion:From 2017 to 2021, the detection capacity of IDD laboratories at the provincial and municipal levels in Shandong Province remains at a high level, and the detection capacity of salt iodine and urine iodine in some county level laboratories still needs to be further improved.