In order to study the pathogenesis of Listeria monocytogenes (LM), we cloned listeriolysin gene into prokaryotic expression vector PET21a. The expression vector was transformed into Escherichia coli BL21 for expression of listeriolysin O (LLO). LLO-His tag fusion protein was purified with a Ni-NTA affinity column and was used as an immunogen to vaccinate BALB/C mice. Hybridomas were developed by fusing mouse myeloma cells Sp2/0 and splenocytes from the immunized mice and screened with purified LLO. Three hybridomas secreting antibodies against listeriolysin O were obtained and named anti-LLO1, anti-LLO2 and anti-LLO3, respectively. Western blotting analysis showed that all of them could specifically bind to the LLO secreted by the LM. The titers of anti-LLO monoclonal antibodies in the supernatants of three hybridomas cultures were 1:3.6 x 10(4), 1:6.4 x 10(4) and 1:1.6 x 10(4), respectively, and the titers of ascites from the hybridoma-injected mice were 1:2 x 10(7), 1:2 x 10(7) and 1:1 x 10(7), respectively, based on ELISA test. The isotypes of the monoclonal antibodies were determined to be IgG1. The dissociation constants (Kd) of these three monoclonal antibodies were determined to be 6.18 x 10(-11), 7.50 x 10(-11) and 6.27 x 10(-11) respectively. These data and reagents will be of great assistance to elucidate the pathogenesis of Listeriosis.
Animals ; Antibodies, Monoclonal ; biosynthesis ; immunology ; Bacterial Toxins ; biosynthesis ; genetics ; immunology ; Cloning, Molecular ; Escherichia coli ; genetics ; metabolism ; Genetic Vectors ; genetics ; Heat-Shock Proteins ; biosynthesis ; genetics ; immunology ; Hemolysin Proteins ; biosynthesis ; genetics ; immunology ; Listeria monocytogenes ; genetics ; pathogenicity ; Mice ; Mice, Inbred BALB C ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; immunology

OBJECTIVETo study the clinicopathological features, immunophenotype, differential diagnosis and prognosis of low-grade nasopharyngeal papillary adenocarcinoma (LGNPPA).

METHODSThe histopathological features and clinical and pathological data of nine cases of LGNPPA were retrospectively analyzed. Immunohistochemistry (Two-step EnVision methods) was used to evaluate the expression of CKpan, vimentin, CK7, CK19, TTF-1 and TG; in situ hybridization was used to detect Epstein-Barr virus mRNA (EBER); and flow-through hybridization was used to evaluate the presence of human papilloma virus (HPV).

RESULTSThe mean age for the nine patients (eight males, one female) was 45.3 years (range 23 to 62 years). Microscopically the tumors were characterized by lobulated, papillary and glandular structures with patchy distribution of spindle cells. The papillary interstitial tissue was edematous, myxoid or hyalinized. The tumors were unencapsulated and infiltrated into the surrounding stroma. Four cases displayed transition between normal nasopharyngeal epithelium to neoplastic cells; and one case contained psammoma bodies. Five cases were strongly positive for CKpan, vimentin, CK7, CK19, TTF-1, and were focally positive for EMA and CD117. These five cases were all negative for TG, CK5/6, CK20, S-100 protein, p63, Calponin and SMA. In situ hybridization for EBER and flow-through hybridization for HPV were negative in all five cases. Follow-up data showed no post-operative recurrence of the LGNPPA.

CONCLUSIONSLGNPPA is a rare low-grade neoplasm with distinct morphological characteristics. Its diagnosis is primarily based on the site of lesions and the histological features. The diagnosis and differential diagnosis of LGNPPA could be aided by immunohistochemical staining. LGNPPA may originate from nasopharyngeal epithelium; and the prognosis is good with simple and complete resection.

Adenocarcinoma, Papillary ; metabolism ; pathology ; Adult ; Carcinoma ; Diagnosis, Differential ; Female ; Herpesvirus 4, Human ; genetics ; Humans ; Immunohistochemistry ; In Situ Hybridization ; Male ; Nasopharyngeal Neoplasms ; metabolism ; pathology ; Neoplasm Proteins ; metabolism ; Nuclear Proteins ; metabolism ; Prognosis ; RNA, Messenger ; metabolism ; Retrospective Studies ; S100 Proteins ; metabolism ; Thyroid Nuclear Factor 1 ; Transcription Factors ; metabolism ; Vimentin ; metabolism

Objective:To investigate the alterations in soft tissue morphology and thickness in the mid-face region of patients with cleft lip and palate (UCLP) secondary to maxillofacial deformity following Le Fort I osteotomy.Methods:A total of 22 patients (16 males and 6 females aged from 17 to 28 years with an average of 20 years) diagnosed with cleft lip and palate secondary to maxillofacial deformity were collected from the Wuhan University Hospital of Stomatology from July 2012 to August 2020. All patients underwent Le Fort I osteotomy. CBCT scans were obtained at T0 (3 days before surgery), T1 (7 days after surgery), and T2 (1 year after surgery). The Dolphin11.95 software and 3D Slicer software were utilized to measure and analyze the soft tissue near the mid-face osteotomy line. Differences in soft tissue thickness before and after surgery were compared.Results:Before and after the operation, the soft tissue thickness at P3, P5, P6, and P8 on the affected side was thicker than that on the healthy side, and the difference was statistically significant, with a P-value of <0.05. At P5, P6, P7, P8, and P9 below the osteotomy line at T2-T0, the degree of postoperative thinning on the affected side was more apparent than that on the healthy side, and there was statistical significance at P6 ( P<0.05). The postoperative soft tissue asymmetry in the Ck region was improved compared with the preoperative one. The preoperative average protruding of the affected side was 0.63 compared with the healthy side, and the postoperative value was 0.17. The preoperative and postoperative Mann-Whitney U tests showed significantly statistical difference. Conclusions:After Le Fort I osteotomy, the facial asymmetry of patients with unilateral cleft lip and palate secondary to maxillofacial deformity is improved. However, there is still a difference in the soft tissue thickness between the healthy side and the affected side, and the change in soft tissue thickness on the affected side is more significant than that on the healthy side.

【Objective】 To investigate the effects of neutrophil-derived exosomes on skin wound healing. 【Method】 ① The venous blood was taken from 10 healthy volunteers to separate and purify neutrophils, and exosomes were prepared from neutrophil supernatants using differential centrifugation on Sepharose columns. ② The rat trauma model with full-thickness skin defect was prepared and randomly divided into three groups: control group, 50 μg/mL exosomes group, and 100 μg/mL exosomes group. The CCK8 test and the scratch test were used to determine the proliferation and migration of vascular endothelial cells, respectively, which was used to investigate the effects of neutrophil-derived exosomes on the formation of endothelial cell tubular structures in vitro. On day 7 and day 14 after surgery, the skin wound healing was observed and the wound healing rate in different treatment groups was compared. On day 14 after surgery, the wound tissues of each group were stained with HE and Masson to observe the arrangement of collagen fibers. 【Results】 ① Neutrophil-derived exosomes were round or oval vesicles with diameter about 100 nm. ② On post injury hour (PIH) 12, the scratch test suggested that wound healing rates in the three groups were close (P>0.05 ), while wound healing rate of 100 μg/mL neutrophil-derived exosomes in the treatment group was higher than that in the control group (P<0.01) and 50 μg/mL exosomes group (P<0.05) on PIH 24. On PIH 12, the result of CCK8 assay showed that 50 and 100 μg/mL exosomes groups had enhanced proliferative activity of vascular endothelial cells (P<0.01) and the results were still higher in both groups than in the control group on PID 24. The results of tube formation assay showed that neutrophil-derived exosomes could promote the formation of tubular structures of endothelial cells in vitro (P<0.01). ③ On day 7 and 14 after surgery, compared with the control group, the wound healing rates of the 50 and 100 μg/mL exosomes groups were higher (P<0.01). The wound healing rate of 100 μg/mL exosomes group was higher than that of 50 μg/mL exosomes group (P<0.01). Compared with the control group, the newly formed epidermis and dermis on the wound surface of the 100 μg/mL exosomes group were longer, the number of collagen fibers increased, and the arrangement direction was highly consistent on the 14 th day after surgery. 【Conclusions】 Neutrophil-derived exosomes can promote full-thickness wound healing in rats by promoting the proliferation and migration of new vascular endothelial cells and the formation of collagen fibrous.

Objective : To analyze the length of hospital stay of patients with heart failure in a multicenter cohort in order to explore the influencing factors of length of stay and provide data support for further intervention. Methods: A total of 2 794 patients enrolled in the multicenter prospective heart failure cohort were divided into two groups: long hospital stay group (≥9 days) and short hospital stay group ( < 9 days) . The general data of the two groups were compared, and the factors with statistical difference in univariate analysis were included in Logistic multifactor regression analysis to explore the difference in length of hospital stay between the two groups. According to left ventricular ejection fraction(LVEF), patients were divided into heart failure with preserved reduced ejection fraction (HFpEF)group, heart failure with mildly reduced ejection fraction(HFmrEF)group and heart failure with reduced ejection fraction(HFrEF) group, and Logistic multifactor regression analysis was performed to find influencing factors. Results : Logistic multifactor regression analysis showed that LVEF, pneumonia, N⁃terminal pro⁃B⁃type natriuretic peptide(NT⁃proBNP), serum sodium, cardiac resynchronization therapy( CRT) or implantable cardioverter defibrillator(ICD)implantation, β blockers, aldosterone receptor antagonists, positive inotropic drugs and vasodilators were all factors influencing the hospitalization of HF patients. In the HFpEF, HFmrEF, and HFrEF groupsCRT/ICD implantation, positive inotonic drugs, and vasodilator use were suggested to be common factors affecting length of hospital stay in all three groups. Conclusion LVEF, pneumonia, NT⁃proBNP, serum sodium, CRT or ICD implantation, β blockers, aldosterone receptor antagonists, positive inotropic drugs and vasodilators are the influencing factors of hospitalization time in HF patients.

OBJECTIVE To explore the effects of posaconazole combined with proton pump inhibitors （PPI） on the blood concentration and the risk of invasive fungal disease （IFD） in patients with malignant hematological disorder. METHODS In accordance with the random number table method， 40 patients with malignant hematological disorders who were admitted to the hematology department of our hospital between December 2020 and December 2021 were chosen and divided into control group （20 cases） and observation group （20 cases）. The control group received Posaconazole oral suspension alone， while the observation group received Posaconazole oral suspension combined with PPI. The incidence of IFD， attainment rate of blood concentration， the time from the start of prophylaxis to IFD onset， the fatality associated with IFD， treatment of infected patients， and blood concentrations of posaconazole on 7th， 14th， 21st， and 28th day after posaconazole application were compared between 2 groups； the occurrence of adverse events during drug administration in the two groups was recorded. RESULTS The study was stopped because 2 patients in the observation group and 9 patients in the control group received hospital departures after taking posaconazole for fewer than 7 days. The incidence of IFD in the observation group was significantly higher than control group， and the attainment rate of blood concentration in the observation group was significantly lower than control group （P＜0.05）. There was no significant difference in the time from the start of prophylaxis to IFD onset， the fatality associated with IFD， treatment of infected patients and the incidence of adverse events （P＞ 0.05）. The blood concentration of posaconazole in the observation group was significantly lower than control group on 7th day of medication （P＜0.05）； there was no significant in blood concentration of posaconazole between 2 groups on the 14th day of medication （P＞0.05）. CONCLUSIONS Posaconazole combined with PPI can reduce the blood concentration of patients with malignant hematological disorders， increase the risk of IFD. Clinical practice should try to avoid the combination of the two or use them under the guidance of therapeutic drug monitoring.

Organ shortage has become one of the major challenges hindering the development of organ transplantation. Xenotransplantation is one of the most valuable methods to resolve global organ shortage. In recent years, the development of genetic engineering technique and research and development of new immunosuppressant have provided novel theoretical basis for xenotransplantation. International scholars have successively carried out researches on xenotransplantation in genetically modified pigs to non-human primates or brain death recipients, making certain substantial progresses. However, most of the researches are still in the preclinical stage, far from clinical application. Therefore, according to the latest preclinical experimental research progress at home and abroad, the history of xenotransplantation, the development of gene modification technology, xenotransplantation rejection and immunosuppression regimens were reviewed, aiming to provide reference for subsequent research of xenotransplantation, promote clinical application of xenotransplantation and bring benefits to more patients with end-stage diseases.