Objective To study the molecular identification method of Sporothrix schenckii based on the nucleotide sequences in internal transcribed spacer region 2 of ribosomal DNA (rDNA) genes. Methods Species-specific primers were used to amplify the internal transcribed spacer region 2 of rDNA of 22 clinical isolates of Sporothrix schenckii and 12 strains of dematiaceous fungi. Totally 11 strains of Sporothrix schenckii were sequenced and analyzed, in which 1 strain came from the US and the others were isolated from different areas in China. A pair of species-specific oligonucleotide primers (SSP) were designed based on the ITS2 sequence. With the species-specific primers, rDNA of Sporothrix schenckii and dematiaceous fungi were amplified by PCR. Results Sequencing and analysis showed that internal transcribed spacer region 2 of rDNA gene was conservative in Sporothrix schenckii. A 300 bp fragment was obtained from 22 strains of Sporothrix schenckii, but not from the other species. Conclusions This method is specific, sensitive and reliable for the identification of Sporothrix schenckii and could be used for clinical molecular diagnosis.

Objective To study the molecular identification method of Candida species and lay the foundation of molecular diagnosis for deep candidiasis. Methods PCR was used to amplify the internal transcribed spacer region of ribosomal DNA from 11 type strains of 9 pathogenic Candida spp. and 39 clinical Candida isolates, and the products were digested with three types of restriction endonucleases (MspⅠ,DdeⅠand HaeⅢ). Results Five patterns of products with different molecular weights were obtained after amplification from 9 Candida spp., and the products were digested with MspⅠ, DdeⅠand HaeⅢ, producing 8, 5 and 4 unique band patterns, respectively. Conclusion PCR-RFLP is reliable, specific and useful for the identification of common pathogenic Candida species.

Purpose To study NF-κB gene expression level in mouse hepatocellular carcinoma cell lines with differently lymphatic metastasis potentials and to discuss its roles in lymphatic metastasis.Methods Using real-time quantitative PCR, NF-κB gene expression level was detected in mouse hepatocellular carcinoma cell lines, including Hca-P with low lymphatic metastasis potential and Hca-F with high lymphatic metastasis potential.Results NF-κB mRNA expression in Hca-P and Hca-F cell lines were (1.41±0.48)×10~(-3),and (2.95±0.22)×10~(-3) (P<0.01),respectively.NF-κB mRNA expression levels were increased with metastasis potential.Conclusion NF-κB gene may play an important role in lymphatic metastasis of hepatocellular carcinoma.

Objective To invstigate the effect of high dose X-ray irradiation on the expression of multidrug resistance-1 (MDR1), Bcl-2, MMP7 genes. Methods A nasopharyngeal carcinoma cell line,CNE1, were irradiated with a total dose of 50 Gy. The resistance to the cisplatin of CNE1 cells and the irradiated CNE1 (CNE1 R) cells was detected by MTT. mRNAs expression of MDR1 , Bcl-2 and MMP7 was measured by quantitative RT-PCR. Results The expression of MDR1 increased in CNE1 R cells. The semiquantitative A value of MDR1 mRNA was 0.47 ±0.04, and the value of CNE1R cells (1st, 7th, 21st,28th, 35th, 42nd and 49th days after irradiation) were 0.67 ± 0. 06 (t = -5.44, P = 0. 003) ,0.70 ± 0. 01(t=-5.90,P=0. 002),0.73±0. 01(t= -6. 45,P=0. 001) ,0. 67 ± 0. 03 (t= -3.97,P=0.011),0.65 ±0.01(t = -4.43,P=0. 007),0. 62±0. 05(t= -2. 64,P=0.046) and 0.62 ±0.02(t = -3.34,P=0.021), respectively. Bcl-2 mRNA expression were 0.55 ±0.02 and 1.05 ±0.04(t = -9.93,P=0. 000) and MMP7 mRNA expression were 0.51 ±0.01 and 0.82 ±0.02(t = -8.51,P=0.000) in CNE1and CNE1 R cells. Conclusions The MDR1 expression was increased after a total dose of50 Gy irradiation,which may be related to the synchronous change of Bcl-2 and MMP7 genes.

OBJECTIVE To investigate the resistant mechanisms of Candida albicans to azoles at molecular level.METHODS NCCLS M-27 protocols were used to test the in vitro susceptibilities of 102 C.albicans strains isolated from the patients with recurrent vulvovaginal candidiasis(RVVC) against fluconazole(FLC) and itraconazole(ITC) to screen the FLC-and ITC-resistant C.albicans isolates;six pairs of primers,A1-A2,B1-B2,C1-C2,D1-D2,E1-E2 and F1-F2 were respectively to amplify gene CYP51 of 4 strains with FLC-and ITC-resistance.The PCR products were sequenced and analyzed to identify the mutation sites by compared with the sequence of gene CYP51 of referenced C.albicans strain in NCBI site of Internet.RESULTS The analysis of full length sequence of CYP51 from 4 FLC-and ITC-resistant strains showed that from total 32 mutation sites there were 4 significant site mutations,where the mutation of GAT to GAC at 116 caused the substitution of D by E(E266D in two strains);GCC to GGT at 117 caused the substitution of A by G(A117G in 1 strain);GAA to GAC at 266 caused the substitution of E by D(E266D in 2 strains);and GTT to ATT at 488 caused the substitution of I by V(V488I in 1 strain).The site mutations of 266 and 488 were tested in 1 strain of 4 strains.CONCLUSIONS The CYP51 total gene of 4 strains has been checked out.Of FLC and ITC-resistant C.albicans alignment in this time,find out 4 significant bp mutations.Causing its amino acide change,among them,A117G has not be interrelated report still now.The details of mechanism need to be further studied.

BACKGROUND: Opiate drugs are widely used to control chronic cancer pain, which brings many adverse reactions. Transplantation of sodium alginate-polylysine-alginate microencapsulated bovine adrenal chromaffin cell (BCC) has reported to be used as chronic cancer pain controllers. However, the high price and poor strength of polylysine limited its clinical application. Chitosan is characterized by sufficient resource, low price and good biocompatibility, which is a substitute for polylysine.OBJECTIVE: To study the effect of xenotransplantation of sodium alginate-chitosan-alginate (ACA) microencapsulated BCC on patients with advanced cancer pain.DESIGN, TIME AND SETTING: A retrospective case analysis. All cases were obtained from Department of Oncology, Affiliated Zhongshan Hospital of Dalian University from January 2007 to December 2008.PARTICIPANTS: Totally 10 patients with advanced cancer, including 1 female and 9 males, aged 46-78 years. According to visual algetic mimic scale (VAS), 3 patients suffered moderate pain and 7 cases suffered severe pain.METHODS: Microencapsulation method was applied to encapsulate BCC with ACA membrane and transplant the microencapsulated BCC (5-7)×10~6 into the subarachnoids pace of 10 patients.MAIN OUTCOME MEASURES: The degree of pain release, duration of analgesic effect, as well as adverse reaction.RESULTS: All 10 patients had pain relief rapidly after transplantation in varying degrees. Complete pain relief was shown in 2 cases, medium relief in 1 case, slight relief in 4 cases. Slight irritation of cauda eguina was presented after transplantation, which could disappear within 3-5 days.CONCLUSION: Xenotransplantation of ACA microencapsulated BCC into the spinal subarachnoids pace of patients with cancer pain can produce analgesic effect promptly, significantly, and safely.

Objective:To analyze the efficacy and safety of radioactive 125I seed implantation in the treatment of unresectable early-stage non-small cell lung cancer (NSCLC), in order to provide data for clinical practice and relevant research. Methods:A retrospective study was conducted on the data of 39 patients with early-stage NSCLC who received CT-guided radioactive 125I seed implantation from Dec 2010 to Dec 2018 in multiple hospitals.The seed implantation process consisted of preoperative planning and design, CT-guided puncture, seed implantation, and postoperative evaluation and dose verification.The efficacy and complications of the treatment were analyzed.The clinical efficacy was evaluated by adopting the Response Evaluation Criteria in Solid Tumors (RECIST) (v1.1) and the adverse reactions were evaluated using the Common Terminology Criteria for Adverse Events (CTCAE v4.0). Results:All the patients were 70 years old on average (51-85). The median lesion diameter was 2.7 cm (1.1-6.0 cm), the median seed activity was 0.7 mCi (0.6-0.8 mCi), while the median follow-up duration was 29 months (3-97 months). Meanwhile, the 1-, 3-, and 5-year overall local control rates were 89.5%, 79%, and 79%, respectively, and the 1-, 3-, and 5-year overall survival rates were 100%, 74.8%, and 49.9%, respectively.Local recurrence and distant metastasis were the main causes of failure, accounting for 17.9% (7 cases) each.The incidence of pneumothorax was 56.4% (22 cases), among which nine cases (23.1%) required invasive closed thoracic drainage.Only 1 case of grade-2 radiation pneumonia (2.6%) was observed, with no other adverse reactions such as dermatoses, esophagitis, or myelitis being discovered.As indicated by univariate analysis, the patients with KPS scores of 80-90, pathological type of adenocarcinoma, T stage of T 1-2, and D90>180 Gy exhibited better local control ( χ2=6.202, P<0.05). Meanwhile, high D90 was also associated with a higher survival rate ( χ2=6.907, P<0.05). Conclusions:Radioactive 125I seed implantation is a safe and effective treatment for unresectable early-stage NSCLC.In cases where external beam radiotherapy is not available, radioactive 125I seed implantation can be considered as one of the treatment options.Pneumothorax is the most common complication of radioactive 125I seed implantation, and adenocarcinoma (pathological type) and higher values of D90 are predictors of better local control.

Objective To obtain a better understaning of the clinical features of Castleman tumor associated paraneoplastic pemphigus. Methods The clinical features and therapy of 10 cases of this disease, diagnosed in the Department of Dermatology of Peking University First Hospital were analyzed. Results Castleman tumor was shown to be the most common neoplasm associated with paraneoplastic pemphigus in China. The clinical presentations, histopathologic characteristics, CT scan findings, and immunologic features were all unique. The early diagnosis and removal of the Castleman tumor are crucial for the treatment of this tumor-associated autoimmune disease. Conclusions Because Castleman tumor is directly related to the induction of autoimmunity, early diagnosis and prompt removal of the tumor are essential to the management of this disease.

Background/Aims: The occurrence and development of hepatitis B virus-associated acute-onchronic liver failure (HBV-ACLF) is closely related to the immune pathway. We explored the heterogeneity of peripheral blood T cell subsets and the characteristics of exhausted T lymphocytes, in an attempt to identify potential therapeutic target molecules for immune dysfunction in ACLF patients. Methods: A total of 83,577 T cells from HBV-ACLF patients and healthy controls were screened for heterogeneity by single-cell RNA sequencing. In addition, exhausted T-lymphocyte subsets were screened to analyze their gene expression profiles, and their developmental trajectories were investigated. Subsequently, the expression of exhausted T cells and their capacity in secreting cytokines (interleukin 2, interferon γ, and tumor necrosis factor α) were validated by flow cytometry. Results: A total of eight stable clusters were identified, among which CD4 + TIGIT + subset and CD8 + LAG-3 + subset, with high expression of exhaust genes, were significantly higher in the HBV-ACLF patients than in normal controls. As shown by pseudotime analysis, T cells experienced a transition from naïve T cells to effector T cells and then exhausted T cells. Flow cytometry confirmed that the CD4 + TIGIT + subset and CD8 + LAG-3 + subset in the peripheral blood of the ACLF patients were significantly higher than those in the healthy controls. Moreover, in vitro cultured CD8 + LAG-3 + T cells were significantly fewer capable of secreting cytokines than CD8 + LAG-3- subset. Conclusions Peripheral blood T cells are heterogeneous in HBV-ACLF. The exhausted T cells markedly increase during the pathogenesis of ACLF, suggesting that T-cell exhaustion is involved in the immune dysfunction of HBV-ACLF patients.

This study was designed to investigate the potential protective effect of isopimpinelline against para-chlorophenylalanine(PCPA)-induced pineal gland damage in rats.Forty male Sprague-Dawley(SD)rats were divided into four groups(n=10 each):a normal group,a model group,a melatonin-treated group(10 mg/kg),and an isopimpinelline-treated group(1.5 mg/kg).All groups,except for the normal,received intraperitoneal injection of PCPA(450 mg/kg)to induce pineal gland damage.Subsequent treatments were administered orally for 7 days.Sleep latency and duration were evaluated on the sixth day using the pentobarbital sodium sleep synergy test.After the treatment period,serum melatonin levels and pineal gland inflammation markers were assessed alongside oxidative and antioxidative parameters.Histological examinations of the pineal gland were conducted,and the expression of proteins related to the Nrf2 and NF-κB signaling pathways were quantified.Data showed that isopimpinelline alleviates the structural damage in the pineal gland of model rats,significantly elevated serum melatonin levels,and markedly improved sleep latency and duration(P＜0.05).Isopimpinelline activated the Nrf2 signaling pathway by inhibiting Keap1 expression,which facilitated the nuclear translocation of Nrf2 and upregulated the antioxidant proteins NQO1 and HO-1,thereby mitigating oxidative stress in the pineal gland(P＜0.05).Furthermore,isopimpinelline significantly reduced the levels of pro-inflammatory cytokines IL-2,TNF-α and IL-6.Isopimpinelline also suppressed the NF-κB signaling pathway,reducing the expression of NF-κB p65,IKKβ,and p-IKKβ proteins,as well as the nuclear translocation of NF-κB p65(P＜0.05),thereby providing anti-inflammatory benefits.In conclusion,isopimpinelline could protect pineal gland from damage by activating the Nrf2 signaling pathway and inhibiting the NF-κB pathway.