Cytotoxic T lymphocytes (CTLs) are critical effectors which play important roles both in anti-tumor and anti-virus immune responses.Through T cell receptors(TCRs),CTLs can specifically recognize MHC- Ⅰ -peptides complexes presented on the surface of target cells,and then release biological substances such as perforin and granzymes into the target cells and dissolve them.Since the significant potential value of CTLs,more and more people are focusing on its basic and appliedstudies.Most researches are about the transformation of the TCR gene.With the development of molecular biology,cloning and transduction of TCR gene have been more mature.Researchers are finding ways to ensure that TCR gene expresses efficiently and assemble correctly.

Objective:To express recombination human interleukin-13(rhIL-13)in methylotropic yeast Kchia pastoris and study its effect on the induction of DC from PBMC in vitro. Methods: An artificial gene with the optional condon usage of Kchia pastoris for IL-13 was designed and synthesized by T4 DNA ligase and PCR.The expression vector pPICZoA-IL-13 was constracted and introduced in Kchia pastoris GS115 by electroporation after linearized with Sacl.Recombinants were selected by plating cells on YPD/Zeocine plates.The recombinant protein secreted by the yeast was identified by 15%SDS-PAGE,ELISA and Western blot.The abilities to induce DC differentiation of IL-13 and IL-4 were compared. Results: Based on the result of the 15% SDS-PAGE and Western blot assay, an about 13 kD recommbinant protein expressed in the yeast supernatant was identified to be recombinant human IL-13. Adherent PBMC cultured in GM-CSF,TNFa and IL-13 displayed morphological characteristic of DC generated in media containing IL-4. They formed cellular clumps and had extensive dendrites sharp. Fluorescence-activated cell sorter analysis showed that they expressed a high level of MHC class II ,CDla,CD80,CD83,CD86. The purity and yield of both DC populations are not significantly different with IL-13 or IL-4. The phagocytosis of immature DC induced by IL-13 is potent. They were also equally potent stimulators of allogeneic lymphocytes in the mixed lymphocyte reaction. Conclusion: rhIL-13 can substitute for IL-4 in the proliferation and development of dendritic cells.

Objective To explore the potential of autologous dendritic cells (DC) pulsed with HLA-A201-binding peptide E613-21(KLPDLCTEL) and E786-94(TLGIVCPI)in inducing specific T cells respouse in vitro.Methods Cervical carcinoma patients with positive HLA-A201 were enrolled and their monocytes isolated and induced into dendritic cells and pulsed with HLA-A201-binding peptide E613-21 and E786-94.PBLs were primed by DCs every week for thee times.The cytokine level of supernatant of CTLs was tested by ELISA.The percentage of special CTLs was tested by flow cytometry.The specific killing effect of CTLs was tested by MTT.Results the numbers of DCs of eleven cervical carcinoma patients were (10.79±0.88) ×106(100 ml peripheral blood).CDllc+HLA-DR+(97.15±2.41)％,CD80+(84.28+5.39)％,CD83 +(85.17±5.06) ％,CD86 + (97.74+0.87) ％.Proliferation index of PBLs primed by DCs three times was 15.4± 1.5.Cytokine levels including IL-2,IL-12,IFN-γ and TNF-α were obviously higher than nonpriming PBLs[(2551.9+195.3) pg/ml,(554.9±64.0) pg/ml,(2416.9±281.7) pg/ml,(632.4 +71.1)pg/ml,respectively] (P＜0.05),but IL-10 was no significant difference between priming CTLs and nonpriming CTLs.The average percentage of special CTLs was obviously higher than control group[(6.32±1.54)％,P＜0.05].The killing effect of CTLs was obviously higher than control group(P＜0.05).Conclusion Dendritic cells pulsed with peptide E613-21 and E786-94 can induce special CTLs in vitro and stimulate CTLs secret cytokines.This will provide science basis for research of therapeutic HPV vaccine.

Objective To investigate the safety and clinical response of dendritic cells (DC)vaccine loaded with HLA-A2-restricted peptides MAGE-3、 Tyr、 MART-1 and GP-100 against malignant melanoma. Methods Twenty three HLA-A2 positive patients with malignant melanoma were enrolled.Peripheral blood mononuclear cells were separated into adherent cells for induction of DC loaded with four peptides. DC vaccine was administered subcutaneously once a week in inguinal region. The immunological responses and clinical responses were evaluated. Results Dendritic cell vaccination were well tolerated in all patients and there was no toxicity observed. Serum levels of IL-2, IL-12 and IFN-γincreased significantly after first vaccination and fourth vaccination. The positive rate of DTH test was 50% (5/10), and 2 fold increase of CD8+ IFN-γ+ cells were observed in 6 of 10 patients. Stable disease was observed in 11 of 23 patients, one patient had a complete metastasis regression, four patients had 50% regression of metastasis,four patients had a minor response, and disease progressed in three patients. Conclusion DC vaccines loaded with peptides MAGE-3、 Tyr、 MART-1 and GP-100 can elicit non-specific and specific immune responses, leading disease control. DC vaccine is considered one of safe and effective treatments for malignant melanoma.

Objective To compared the reliability and validity of the application of two consciousness assessment scales in neurosurgical patients by Full Outline of Unresponsiveness Score Coma Scale (FOUR) and the Glasgow Coma Scale (GCS), and provide the effective evaluation for the consciousness of the nerve severe patients. Methods A total of 100 neurological intensive patients from Nanfang Hospital, Southern Medical University were enrolled and the consciousness was evaluated by FOUR and GCS. The reliability and validity of these scales were evaluated and compared by the following method such as Cronbach αcoefficient, the inter-rater agreement,content validity index (CVI) and the receiver operating characteristic curve (ROC) which used to predict the discrimination of prognosis.The important experimental indexes of blood of the degree of brain injury were collected in the same period:S-100 B protein and neuron specific enolization enzymes (NSE). Results The Cronbach α coefficient was 0.811 (FOUR) and 0.923(GCS). The overall rater agreement was excellent with an intraclass correlation coefficients of 0.972 (FOUR) and 0.979(GCS). CVI was 0.965 (FOUR) and 0.973 (GCS). Both scales had better distinguish and predictive abilities for the poor prognosis.The area under the curve for mortality was 0.938 for the FOUR and 0.932 for the GCS. The best cut-off values for predicting poor prognosis were FOUR of 12 and GCS of 11. For the FOUR, the correlation coefficient was-0.324(P<0.05) with the level of NSE,-0.427(P<0.01) with the level of S-100B protein. For the GCS,the correlation coefficient was-0.316 (P<0.05) with the level of NSE,-0.395 (P<0.01) with the level of S-100B protein. Conclusions Both FOUR and GCS are a reliable scale for evaluating the level of consciousness in neurosurgical patients. The GCS is familiar with the medical staff while the FOUR is more adapted to assess the patients with tracheotomy or intubated and which more easy to remember and learn by medical staff of neurosurgery department.

OBJECTIVETo explore the effect of recombinant human soluble CD(40) ligand (rhsCD(40)L) and CD(40)L cDNA transfected cell (CD(40)L-TC) on the behavior of malignant B lymphocytes, and investigate the possibility of using rhsCD(40)L as a new bio-factor in tumor immunotherapy.

METHODrhsCD(40)L and CD(40)L-TC were obtained by gene recombinant techniques. Multiple myeloma cell lines, XG2, XG7, U266 and 8226, B-lymphoma cell lines, Raji and Daudi were selected to detect responses to rhsCD(40)L and CD(40)L-TC stimulation. Cell growth curve, cell cycle, early apoptosis as well as membrane surface molecules on these cell lines were analyzed.

RESULTS(1) The expression levels of CD(40) molecule on malignant B lymphocytes showed heterogeneity. High level of CD(40) on XG2, moderate on 8266, Raji, and Daudi, and no expression on U266 and XG7 were detected. The rhsCD(40)L stimulation gave rise to a typical homo-type cell aggregation of XG2 and Daudi. Meanwhile, at least 10 to 20 of CD(40)(+) XG2 or CD(40)(+) Daudi cells were found adherent to one pre-treat ed CD(40)L-TC. (2) Co-incubation with rhsCD(40)L (5 micro g/ml), or CD(40)L-TC (tumor cell: CD(40) = 5:1) resulted in a significant inhibition of in vitro cell growth of XG2, Raji and Daudi, with G(1)-phase arrest for XG2 and G(2)-phase for Raji and Daudi. These two kinds of CD(40) stimulators induced XG2, Raji and Daudi cells to apoptosis in vitro. The apoptotic rate for XG2 was 23.3% (rhsCD(40)L) and 18.8% (CD(40)L-TC), for Daudi 14.2% and 15.9%, and for Raji 11.6% and 8.9% respectively. (3) Phenotype analysis showed that CD(95) expression levels were significantly up-regulated on XG2, Raji and Daudi after stimulation with rhsCD(40)L or CD(40)L-TC, and CD(80) and CD(18) expression levels on Raji were respectively enhanced and decreased.

CONCLUSIONThe abilities to directly inhibit XG2, Daudi and Raji cell proliferation, to induce themapoptosis, as well as to up-regulate immune co-stimulator molecule CD(80) expression on Raji cells would make rhsCD(40)L a potential bio-factor for tumor immuno-therapy.

B-Lymphocytes ; drug effects ; metabolism ; pathology ; CD40 Antigens ; metabolism ; CD40 Ligand ; genetics ; metabolism ; pharmacology ; Cell Division ; drug effects ; Coculture Techniques ; DNA, Complementary ; genetics ; Humans ; Lymphoma, B-Cell ; metabolism ; pathology ; Recombinant Proteins ; pharmacology ; Time Factors ; Transfection ; Tumor Cells, Cultured ; drug effects

Estrogen is an important hormone secreted by the female reproductive system. Its main function is associated with reproduction, growth and development. Studies have shown that estrogen has biological functions such as regulating vasoconstriction, antioxidant stress, anti-inflammatory and neuroprotection, and also affects brain structure and network. Studies have shown that estrogen is closely associated with the occurrence and development of white matter hyperintensities (WMHs). This article reviews the relationship between estrogen and menopausal hormone replacement therapy and WMHs, and their possible pathophysiological mechanisms.

Carotid atherosclerosis (CAS) is closely associated with the decline of cognitive function in the elderly, which can lead to persistent or progressive cognitive function and neurological dysfunction. Vascular cognitive impairment (VCI) is considered to be an intervenable disease. Studies have shown that CAS is one of the main causes of VCI. Further study on the relationship between CAS and VCI will help to better prevention and treatment of VCI.

Neutrophil to lymphocyte ratio (NLR) can provide information of local or systemic inflammation and immune status. With the increasing attention to the role of inflammatory and immune factors in vascular cognitive impairment (VCI), it is very important to find new serum inflammatory markers for early identification and intervention of VCI. This article reviews the related research on NLR, VCI and their risk factors, expounds the role of inflammation in the pathogenesis of VCI, and provides help for the diagnosis and prevention of VCI.

In industrial fermentation processes, bacteria have to adapt environmental stresses. Sometimes, such a self-adaption does not work and will cause fermentation failures, although such adaptation also can generate unexpected positive effects with improved fermentation performance. Our review introduces cell self-adaption to environmental variations or stress, process optimization based on such self-adaptions, with heterologous proteins production by Pichia pastoris and butanol fermentation as examples. Our review can sever as reference for fermentation optimization based on cell self-adaption.
Adaptation, Physiological ; Butanols ; metabolism ; Environment ; Fermentation ; Pichia ; cytology ; metabolism