Preparation of mouse monoclonal antibodies against human adenovirus 55 Hexon (HAdV55 Hexon) protein / 细胞与分子免疫学杂志

Objective To prepare specific mouse monoclonal antibody (mAb) against human adenovirus type 55 Hexon protein (HAdV55 Hexon). Methods The Hexon genes of HAdV55, 3, 4, 7, 16 and 21 were chemically synthesized as templates for PCR amplification. The prokaryotic expression plasmids pET28a-HAdV55 Hexon and eukaryotic expression plasmids pCAGGS-HAdV3, 4, 7, 16, 21 and 55 Hexon were constructed respectively. The pET28a-HAdV55 Hexon plasmid was transformed into E. coli competent cell BL21 (DE3) and was induced by IPTG. After the purified inclusion body was denatured and renatured, Hexon55 protein was purified by tangential flow filtration system. pCAGGS-HAdV55 Hexon was used to immunize BALB/c mice by cupping, and HAdV55 Hexon protein was used to booster immunization. The anti-HAdV55 Hexon mAb was prepared by hybridoma technique and the titer and subclass were determined. The specificity of antibody was identified by Western blot using HEK293T cells transfected with pCAGGS-HAdV55 Hexon and by immunofluorescence assay (IFA) using BHK cells transfected with pCAGGS-HAdV55 Hexon. Both clones with high titer were selected, and the cross-reactivity of pCAGGS-HAdV3, 4, 7, 16, 21 and 55 Hexon transfected cells were analyzed by Western blot analysis and IFA. Results PET28a-HAdV55 Hexon and pCAGGS-HAdV55 Hexon, 3, 4, 7, 16 and 21 expression plasmids were successfully constructed. BL21 transformed with pET28a-HAdV55 Hexon was induced by IPTG. The HAdV55 Hexon protein was mainly expressed in the form of inclusion body. After denaturation and renaturation, the purified HAdV55 Hexon protein was obtained by ultrafiltration. Six hybridoma cell lines secreting HAdV55 Hexon mAb were obtained. The antibody subclass analysis showed that 2 strains were IgG2a subtypes and 4 strains were IgG2b. Two specific HAdV55 Hexon antibodies with high titer were obtained, and there was no cross-reactivity with HAdV3, 4, 7, 16, 21 Hexon. Conclusion The specific mice mAb against HAdV55 Hexon provides an experimental basis for establishing its antigen detection method.

Effects of FAC combined with Bu conditioning for SAA patients undergoing allogeneic stem cell transplantation / 西安交通大学学报(医学版)

【Objective】 To observe the effects of FAC combined with Bu conditioning for severe aplastic anemia (SAA) patients undergoing allogeneic hematopoietic stem cell transplantation. 【Methods】 The data of 28 SAA patients who underwent allogeneic hematopoietic stem cell transplantation in our hospital from January 2016 to September 2021 were collected and analyzed. The patients were divided into two groups: FAC conditioning group and FAC + Bu conditioning group. We observed the side effects of conditioning regimen, hematopoietic recovery, acute and chronic graft versus host disease (GVHD), viral infection, incidence of venous obstructive disease (VOD), progression free survival (PFS), and overall survival (OS). 【Results】 There was no significant difference between the two groups in age, gender, physical condition, history of disease, gender relationship between donors and recipients, transplantation type, infusion of bone marrow nucleated cells, MNC or CD34 positive cells (P>0.05). Compared with those in FAC group, the side effects of auditory hallucination and visual hallucination in FAC + Bu group increased, the incidence of mixed chimerism decreased, the time of platelet reconstruction shortened, the incidence of grade Ⅲ-Ⅳ aGVHD increased. However, 3-year PFS or OS did not significantly differ between the two groups. 【Conclusion】 FAC combined with Bu conditioning regimen promotes implantation and reduces mixed chimerism. However, it does not improve patients’ overall survival.

Epidemiological characteristics and clinical features of visceral leishmaniasis in 41 children / 中华传染病杂志

Objective:To investigate the epidemiological and clinical characteristics of visceral leishmaniasis (VL) in children, and to analyze the distinguishing features of VL associated hemophagocytic lymphohistiocytosis (HLH), so that to provide reference for the diagnosis and treatment of VL.Methods:Forty-one children with VL admitted to Xi′an Children′s Hospital from July 2012 to June 2021 were enrolled. The clinical data were retrospectively analyzed, including epidemiology, clinical manifestations, laboratory data, diagnostic methods, treatment regimens and outcomes. The patients were divided into VL group and VL+ HLH group according to whether combined with HLH or not, and the clinical characteristics and laboratory findings of the two groups were compared. Two independent samples t test, Mann-Whitney U test and chi-square test were used for statistical analysis. Results:Forty-one children with VL were from different provinces, including Shaanxi Province (70.73%(29/41)), Gansu Province (14.63%(6/41)), Shanxi Province (12.20%(5/41)) and Ningxia Hui Autonomous Region (2.44%(1/41)), and 87.80%(36/41) of them lived in rural areas. The peak age was >1.0 to 3.0 years old (63.41%(26/41)). They were sporadic throughout the year. The main clinical manifestations included fever (97.56%(40/41)), splenomegaly (95.12%(39/41)), lymphadenopathy (82.93%(34/41)) and hepatomegaly (60.98%(25/41)). The numbers of cases that Leishman-Donovan bodies were detected in the first, second and third bone marrow smears were 36, four and one, respectively. Anemia, thrombocytopenia and leukopenia detected by blood routine test were 100.00%(41/41), 78.05%(32/41) and 58.54%(24/41), respectively. There were statistically significant differences in the platelet count, lactate dehydrogenase, alanine aminotransferase, triglycerides, fibrinogen and ferritin between VL group (28 cases) and VL+ HLH group (13 cases) ( t=-2.56, t=2.64, Z=-2.66, t=7.15, t=-5.76 and t=3.86, respectively, all P<0.050). The proportions of hepatomegaly and hemophagocytes found in the bone marrow smears in VL group were both lower than those in VL+ HLH group, and the differences were both statistically significant ( χ2=4.47 and 10.93, respectively, both P<0.050). Twelve cases with VL+ HLH were treated with antimony (for six days) and intravenous immunoglobulin, and the others were treated with antimony only. The cure rates of the patients treated with antimony for one and two courses were 92.68%(38/41) and 4.88%(2/41), respectively. The dose of antimony was increased one third and treatment course was prolonged to eight days in one cured case. After (41.36±31.49) months of follow-up, three cases recurred after five to eight months of cure and all of them were cured after one more course of treatment with antimony. Conclusions:Children with VL are mainly distributed in rural areas. The common clinical manifestations are fever and involvement of reticuloendothelial system, which are not specific. The positive rate of Leishman-Donovan bodies found in bone marrow smears is high, and a few negative cases need repeated bone marrow aspiration. Standardized treatment with antimony for VL in children is effective, and combination therapy with immunoglobulin can be considered if patients with VL associated HLH. Very few cases may recur and antimony is still effective.

Strategies for engineering the thermo-stability of glycosidase / 生物工程学报

Glycosidases are widely used in food and pharmaceutical industries due to its ability to hydrolyze the glycosidic bonds of various sugar-containing compounds including glycosides, oligosaccharides and polysaccharides to generate derivatives with important physiological and pharmacological activity. While glycosidases often need to be used under high temperature to improve reaction efficiency and reduce contamination, most glycosidases are mesophilic enzymes with low activity under industrial production conditions. It is therefore critical to improve the thermo-stability of glycosidases. This review summarizes the recent advances achieved in engineering the thermo-stability of glycosidases using strategies such as directed evolution, rational design and semi-rational design. We also compared the pros and cons of various techniques and discussed the future prospects in this area.

Clinical features and genetic testing of a Chinese pedigree affected with Smith-Lemli-Opitz syndrome / 中华医学遗传学杂志

OBJECTIVE@#To analyze the clinical features and genetic variants of two patients from a pedigree affected with Smith-Lemli-Opitz syndrome and explore their genotype-phenotype correlation.@*METHODS@#Clinical data and family history of the pedigree were collected. Whole exome sequencing was carried out to identify the potential variants. Suspected variants were verified by Sanger sequencing of the family members.@*RESULTS@#The proband and her sister both presented with feeding difficulty, facial dysmorphism, seizures, and mental and speech retardation. The third child of this family presented with feeding difficulty, poor weight gain and severe malnutrition after birth. He had died of unknown cause at 6 months without genetic testing. The fourth child was a healthy boy. Genetic testing showed that both the proband and her sister have carried c.127G>T (p.Val43Phe) and c.820_825del (p.Asn274_Val275del) compound heterozygous variants of the DHCR7 gene (NM_001360.2), but the fourth child carried neither of the variants. The two variants were unreported in the literature and disease-related databases, and were not included in the 1000G and gnomAD databases. The c.820_825del variant may affect the sterol-sensitive region of the DHCR7 protein, which can lead to deletion of two amino acids at positions 247 and 275, causing truncation of the DHCR7 protein. It is speculated that this may affect the stability of protein's spatial conformation, thereby decrease the activity of the enzyme. The c.127G>T variant may affect the first transmembrane region of the protein, which is involved in the transmembrane transport of proteins. Multiple software predicted it to be harmful. Conservation analysis suggested that the three amino acids all locate in a highly conserved region of the protein. In consideration of the clinical phenotype, family history and result of genetic testing, we speculated that both patients had Smith-Lemli-Opitz syndrome due to variants of the DHCR7 gene.@*CONCLUSION@#This pedigree has enriched the phenotypic and genotypic data of Smith-Lemli-Opitz syndrome, which clarified the genetic etiology of the patients and provided a basis for genetic counseling of this pedigree.

Therapeutic effect comparison of modified Miccoli operation and conventional open thyroidectomy in the treatment of benign thyroid disease / 国际外科学杂志

Objective:To compare the efficacy of traditional open thyroidectomy and modified Miccoli in the treatment of benign thyroid diseases.Methods:The data of 343 patients with benign thyroid disease in Xi′an Central Hospital and Xi′an Third Hospital were retrospectively analyzed. According to the surgical method, the patients were divided into the observation group (modified Miccoli operation, n=169) and the control group (traditional open operation, n=174). The operation effect, postoperative recovery, patient satisfaction, postoperative complications and recurrence within 1 year of follow-up were compared between the two groups.Count data were represented by n(%), measurement data consistent with normal distribution were represented by Mean± SD, and non-normal data were represented by M( P25, P75). T test or Wilcoxon rank sum test were used to compare the measurement data between groups according to their distribution characteristics. Comparison of statistical data between groups was made by chi-square test. Results:The intraoperative blood loss and postoperative drainage volume of the patients in the observation group were 26.2 (6.9, 40.8) ml and (33.4±8.6) ml, respectively, which were significantly less than those in the control group 50.6 (37.5, 63.7) ml and (35.7±9.5) ml (all P<0.05). The postoperative recovery time in the observation group was (3.9±2.8) d, which was significantly shorter than that in the control group (5.1±2.4) d ( P=0.001). Postoperative complications such as hoarseness, subcutaneous ecchymosis, scar formation and decreased parathyroid function were significantly different between the two groups ( P<0.05). The recurrence rate of 7.69% in the observation group was significantly lower than that of 19.54% in the control group 12 months after operation ( P<0.05). Conclusions:The modified Miccoli in the treatment of benign thyroid diseases not only has the advantages of less trauma, faster postoperative recovery, less pain and higher patient satisfaction, but also has lower postoperative complications and recurrence rate, which is worthy of clinical promotion and application.

Clinical Study on Oral Administration and External Application of Yanyuxiao on Acute Traumatic Synovitis of Knee Joint / 中国中医药信息杂志

Objective To observe the clinical efficacy of oral administration and external application of Yanyuxiao on acute traumatic synovitis of knee joint. Methods Totally 86 patients with acute traumatic synovitis of knee joint were randomly divided into treatment group and control group, with 43 cases in each group. The patients in the two groups were treated with conventional western medicine. Patients in the control group orally took diclofenac sodium at the same time, while patients in the treatment group received oral administration and external application of Yanyuxiao Formula for 2 weeks. The scores of pain, swelling and function of knee joint and the levels interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) of joint fluid before and after the treatment were observed. The efficacy and the recurrence rate in three months after treatment were compared. Results The scores of pain, swelling and function of knee joint of the two groups significantly decreased after treatment, and the scores of pain, swelling and function of knee joint of the treatment group in one and two weeks were lower than those of the control group (P<0.05, P<0.01). The levels of synovial fluid IL-6 and TNF-α of the treatment group were lower than those of the control group after two-week treatment (P<0.01); the total effective rate was higher than that of the control group (P<0.05). The recurrence rate of the treatment group was lower than that of the control group after discharged for 3 months (P<0.05). There were no obvious adverse reactions during the treatment period. Conclusion Oral administration and external application of Yanyuxiao Formula has definite efficacy for acute traumatic synovitis of knee joint, with good safety.

Effect of cholecystokinin on intestinal interstitial cells of Cajal and small intestinal motor function in acute pancreatitis rats / 中华临床营养杂志

Objective To investigate the effect of cholecystokinin (CCK) on rat intestinal interstitial cells of Cajal (ICC) and the small intestinal motor function during experimentally induced acute pancreatitis (AP).Methods 20 male SD rats were divided with a random number table into model group (n =15) and control group (n =5):the 15 rats in the model group were further divided into 3 subgroups,i.e.0,20,40 μg/kg CCK sub-groups.L-omithine was intraperitoneally injected to induce pancreatitis,while normal saline was injected in the control group.24 hours after establishing AP model,CCK was administered intraperitoneally every day for 5 consecutive days.The rats were sacrificed by cervical dislocation 30 minutes after gavage with 2 ml ink.Length of the small intestine which have become black and that of the whole small intestine were measured.Intestinal sections were studied by confocal microscopy after immunofluorescence with specific antibodies.The number of positive cells was compared among the groups.Results In the control group,the number of ICC in the myenteric plexus and deep muscular plexus was (21.16 ± 3.19) /field and (17.20 ± 1.75) /field,respectively;in the AP group was (5.00 ± 1.05) /field and (4.52 ± 1.05) /field,respectively;in the AP + CCK20 group was (10.76 ± 2.09) /field and (9.84 ± 1.68) /field,respectively;in the AP + CCK40 group was (13.72 ±2.97) /field and (12.40 ± 1.81) /field,respectively;the number of ICC in the AP group were significantly lower than those in the control group (P =0.001).The ratio of transmission distance in small intestine in the control,AP,AP + CCK20,and AP + CCK40 groups was 0.71 ± 0.05,0.54 ± 0.07,0.68 ± 0.10,and 0.74 ± 0.07,respectively.The intestinal motor function was reduced in the AP group,demonstrated by shortened transmission distance,which was improved after the administration of CCK (P =0.003).Conclusions Pancreatitis can cause ICC damage,reduce ICC number,destroy ICC network and the small intestinal motor function.CCK can mitigate the damage to ICC and protect the integrity of ICC network,thereby improving intestinal motor function.

Curative observation of Huoxue-Zhitong decoction combined with mannitol in the treatment of swelling and pain after operation of tibiofibular fracture / 国际中医中药杂志

Objective To observe the clinical curative effect of Huoxue-Zhitong decoction combined with mannitol in the treatment of swelling and pain after operation of tibiofibular fracture. Methods 86 patients with swelling after the operation of fracture of tibia and fibula were randomly divided into two groups, with 43 cases in each group. The observation group was treated with intravenous rapid infusion of mannitol, the control group was treated with Huoxue-Zhitong decoction combined with intravenous rapid infusion of mannitol. Swelling, pain visual analogue scale (VAS) score of limb of patients at 1 D and 10 d after operation, time of detumescence and acesodyne and the total efficiency of the treatment between the two groups were compared. Results The indices of the two groups mentioned above at 7 d after the operation significantly decreased, the value in the observation group was lower than the control group (1.3 ± 0.7 cm vs. 1.9 ± 0.9 cm, t=3.451; 1.5 ± 0.6 points vs. 1.9 ± 0.7 points, t=2.845; all P<0.01); The detumescence and acesodyne time of the observation group was shorter than those of the control group (8.6 ± 1.7 d vs. 9.8 ± 2.3 d, t=2.751; 5.5 ± 1.4 d vs. 6.7 ± 1.7 d, t=3.573; all P<0.01), the total efficiency in the observation group was superior to the control group, there were statistically significant differences (97.7% vs. 83.7%; χ2=4.961, P=0.026). Conclusions Huoxue-Zhitong decoction combined with mannitol is effective and safe in the treatment of swelling and pain after operation of tibiofibular fracture.

Prokaryotic expression and biological functions of Mycobacterium tuberculosis Rv1884c and Rv0867c genes / 中华检验医学杂志

Objective To construct fusion gene and prokaryofie expression plasmid encoding the Rv1884c and Rv0867c genes in Mycobacterium tuberculosis (M.tb).The fusion protein wsg expressed efficienfly in E.coli cells.Methods The Rv1884c and Rv0867c genes were amplified by polymerase chain reactions (PCR) with specific primers from genomic DNA of M.tb H37Rv strain,and cloned into pGEX-4Tland pUC19 vectors.Rv1884c and Rv0867c were subcloned into the expression vector pGEX-4T-1 and pPRO-EXHT followed by DNA sequencing.The plasmids were transformed into E.coli DH5α and induced to produce GST-fused Rv1884c and His-fused Rv0867c fusion protein.The protein molecular weight and expression format was analyzed by SDS-PAGE.Results The recombinant expressive vectors pGEX-4T-1Rv1884c and pPRO-EXHT-Rv0867c were constructed.The DH5α strains of E.coli with recombinant plasmid showed high level of Rv1884c and Rv0867c gene expressions after IPTG induction.The SDS-PAGE showed that the plasmids expressed Rv1884c and Rv0867c fusion proteins with molecule weight of 45 000 and 80 000.The recombinant protein accounted for 18.3%and 23.7%of total bacteria protein.The expressed proteins could be purified via GSTrao FF and Ni2+-NTA system kits in denatured condition.Conclusions Mycobacterium tuberculosis Rv1884e and Rv0867c genes have been cloned and expressed Successfully in E.coli DH5α.The results lay a basis for further study of fast culfivation in Mycobacterium tuberculosis and investigation of their activities and functions.