Recent advances in simultaneous detection strategies for multi-mycotoxins in foods.

Mycotoxin contamination has become a challenge in the field of food safety testing, given the increasing emphasis on food safety in recent years. Mycotoxins are widely distributed, in heavily polluted areas. Food contamination with these toxins is difficult to prevent and control. Mycotoxins, as are small-molecule toxic metabolites produced by several species belonging to the genera Aspergillus, Fusarium, and Penicillium growing in food. They are considered teratogenic, carcinogenic, and mutagenic to humans and animals. Food systems are often simultaneously contaminated with multiple mycotoxins. Due to the additive or synergistic toxicological effects caused by the co-existence of multiple mycotoxins, their individual detection requires reliable, accurate, and high-throughput techniques. Currently available, methods for the detection of multiple mycotoxins are mainly based on chromatography, spectroscopy (colorimetry, fluorescence, and surface-enhanced Raman scattering), and electrochemistry. This review provides a comprehensive overview of advances in the multiple detection methods of mycotoxins during the recent 5 years. The principles and features of these techniques are described. The practical applications and challenges associated with assays for multiple detection methods of mycotoxins are summarized. The potential for future development and application is discussed in an effort, to provide standards of references for further research.

Effect of ultrasonic vibration on forming force and forming quality in micro-punching with a flexible punch.

In this paper, a special ultrasonic microforming method, Micro Ultrasonic thin Sheetmetal Forming using molten plastic as a flexible punch (short as Micro-USF), was used to conduct micro-punching experiments on stainless steel sheet with thickness of 10 µm and 20 µm. The influence of ultrasonic vibration on forming force and forming quality were investigated. The experimental results showed that the forming force required for punching thin sheet metal decreased gradually as the ultrasonic time or ultrasonic power increased. By applying the ultrasonic vibration effect, the forming force could be decreased dramatically and the maximum value of forming force drop could reach 86%. Moreover, with the application of ultrasonic vibration, the size accuracy and shape accuracy of micro-holes could be increased by 36.92% and 22.65%, but the cross-section quality of micro-holes were not significantly improved.

Gata2-L359V impairs primitive and definitive hematopoiesis and blocks cell differentiation in murine chronic myelogenous leukemia model.

GATA2, a key transcription factor in hematopoiesis, is frequently mutated in hematopoietic malignancies. How the GATA2 mutants contribute to hematopoiesis and malignant transformation remains largely unexplored. Here, we report that Gata2-L359V mutation impeded hematopoietic differentiation in murine embryonic and adult hematopoiesis and blocked murine chronic myeloid leukemia (CML) cell differentiation. We established a Gata2-L359V knockin mouse model in which the homozygous Gata2-L359V mutation caused major defects in primitive erythropoiesis with an accumulation of erythroid precursors and severe anemia, leading to embryonic lethality around E11.5. During adult life, the Gata2-L359V heterozygous mice exhibited a notable decrease in bone marrow (BM) recovery under stress induction with cytotoxic drug 5-fluorouracil. Using RNA sequencing, it was revealed that homozygous Gata2-L359V suppressed genes related to embryonic hematopoiesis in yolk sac, while heterozygous Gata2-L359V dysregulated genes related to cell cycle and proliferation in BM Lin-Sca1+c-kit+ cells. Furthermore, through chromatin immunoprecipitation sequencing and transactivation experiments, we found that this mutation enhanced the DNA-binding capacity and transcriptional activities of Gata2, which was likely associated with the altered expression of some essential genes during embryonic and adult hematopoiesis. In mice model harboring BCR/ABL, single-cell RNA-sequencing demonstrated that Gata2-L359V induced additional gene expression profile abnormalities and partially affected cell differentiation at the early stage of myelomonocytic lineage, evidenced by the increase of granulocyte-monocyte progenitors and monocytosis. Taken together, our study unveiled that Gata2-L359V mutation induces defective hematopoietic development and blocks the differentiation of CML cells.

In situ resuspension rate monitoring method in the littoral zone with multi-ecotypes of a shallow wind-disturbed lake.

Sediment resuspension has been recognized as a crucial internal process in aquatic ecosystems. However, there is still a lack of reliable measuring methods due to the complex hydrodynamic conditions in large shallow eutrophic lakes. In this study, sequential sediment traps (SST) and instantaneous multiple point (IMP) methods were compared at 6 sites located in the littoral zone of Zhushan Bay in Lake Taihu. Results show that the average resuspension rates (RRs) estimated using the IMP method at sites 1 to 6 were 266.39, 272.79, 235.17, 254.95, 392.25, and 483.85 g·m-2d-1, respectively. While the RRs estimated using the SST method were 195.16, 236.99, 116.76, 156.23, 389.53, and 509.85 g·m-2d-1, respectively. In wind-disturbed areas, both methods were suitable for RR analysis in large and shallow eutrophic lakes and SST provides high-resolution temporal RR estimations. However, in the areas with cyanobacterial blooms and vegetation cover, the IMP method overestimated the RR. Therefore, SST was more suitable across different conditions in large and shallow eutrophic lakes, providing a simple, accurate, and high-resolution temporal estimation of RR, while furthering our understanding of lake evolution processes.

[Effect of Nutrient Loadings on the Regulation of Water Nitrogen and Phosphorus by Vallisneria natans and Its Photosynthetic Fluorescence Characteristics].

Submerged macrophytes are an important component of aquatic ecosystems. During the growing period, submerged macrophytes can absorb nitrogen and phosphorus nutrients to reduce pollution loadings. Shoots of submerged macrophytes can also promote the adhesion of suspended substances in water, reducing the turbidity. The release of nutrients in sediments can be suppressed by its root system, and the resuspension of sediments caused by disturbance of winds and waves can also be resisted. The role of submerged macrophytes in ecological restoration of eutrophic lakes has attracted widespread attention. In 1960, the submerged plants Vallisneria natans and Potamogeton malaianus had been the dominant species in East Taihu. However after 2002, Nymphoides peltatum, Elodea nattalii, P. malaianus, etc. have gradually taken over the dominant roles along with significant elevations of nitrogen and phosphorus levels. Nutrients in water are not the only key factors causing eutrophication of water bodies; the nutrient source for submerged plant growth affect both the purification efficiency and the photosynthetic characteristics of submerged macrophytes. Excessive nitrogen and phosphorus concentrations can inhibit the photosynthetic physiological activities of submerged macrophytes, affecting the succession of aquatic vegetation. In addition, under high nutrient conditions, the competition from periphytic algae and planktonic algae may also directly poison submerged macrophytes, leading to its degradation and disappearance. Systematic studies on the regulation and photosynthetic fluorescence response mechanism of submerged macrophytes to varied nutrient loadings are helpful in revealing their relationships. The seedlings of submerged macrophyte V. natans were transplanted in a laboratory mesocosm to study the effect of nutrient loadings on its regulation of water nitrogen and phosphorus. Three nitrogen and phosphorus loadings from low, medium, and high levels derived from nitrate, ammonium, and phosphate were setup as the aquatic medium for the plant growth. Twelve harvests were carried out to determine the evolution of nutrient removal performance of V. natans. Its photosynthetic fluorescence characteristics were measured by a pulse-amplitude modulated fluorometer (Diving-PAM). Results showed that the nitrogen and phosphorus adsorption abilities of V. natans were gradually enhanced with the increase of nutrient concentrations in the range of TN ≤ 12 mg·L-1 and TP ≤ 1.0 mg·L-1. In the treatment of high nutrient concentrations (TN=12 mg·L-1 and TP=1.0 mg·L-1), the removal rates of nitrogen and phosphorus reached more than 95%. V. natans preferentially absorbed ammonium nitrogen when its concentration was high. The medium nutrient concentrations (TN:8-12 mg·L-1 and TP:0.6-1.0 mg·L-1) did not significantly affect the Fv/Fm ratio of leaves. However, the low nutrient concentrations (TN=3 mg·L-1 and TP=0.3 mg·L-1) could improve the Fv/Fm ratio of leaves and were beneficial for the growth of V. natans. The inhibition of photosynthetic activity and light tolerance were enhanced with the increase in nutrient concentrations. The photosynthetic activity of V. natans gradually recovered with no significant changes in the capacity for light harvesting, when the nutrient concentrations gradually decreased in the water. Our results indicate that the high nitrogen and phosphorus loadings indeed hamper the photosynthetic capacity, which may subsequently restrain the maintenance of the dominance of V. natans in the submerged macrophyte communities.

[Deposition Characteristics of Suspended Solids and the Response of Dissolved Nutrients in Spring in the Western Lakeside of Taihu Lake].

To reveal the spatiotemporal characteristics of nutrients in the deposition process of suspended solids in lakeside zone,in situ deposition tests were performed in the western lakeside of Taihu Lake,and the contents of TP,TN,NH4+-N and NO3--N were measured and analyzed.The results showed that the deposition fluxes in the western lakeside of Taihu Lake ranked as follows:artificial reed areas >non-vegetation nearshore areas >natural reed areas >non-vegetation offshore areas,with their average values of (1383.40±925.60),(1208.67±743.50),(278.72±142.53),(245.58±154.25) g·(m2·d)-1,respectively.From the 6th day,the deposition volume steadily increased,with the deposition rate larger than the decomposition rate.Through the 15-day continuous in situ observation,the content of TP in nearshore zone was 2-3 folds larger than that of offshore zone,and the content of NH4+-N was significantly different from that of NO3--N in the settlement bottle (P<0.01).The deposition volume was significantly and positively correlated to both TN and NH4+-N contents in the water column (P<0.01,n=42),suggesting that the TN and NH4+-N contents in the overlying water increased with the deposition fluxes.The correlation coefficient between TN and NH4+-N was 0.84,implicating that the increase of deposition flux may accelerate the mutual transformation between different forms of nitrogen.These findings should be taken into account in the current control of black blooms and nutrient management in Taihu Lake.

Des-aspartate-angiotensin I causes specific release of PGE2 and PGI2 in HUVEC via the angiotensin AT1 receptor and biased agonism.

DAA-I (des-aspartate-angiotensin I), an endogenous angiotensin, had been shown earlier to ameliorate animal models of cardiovascular diseases via the angiotensin AT1 receptor and prostaglandins. The present study investigated further the action of DAA-I on the release of PGE2, PGI2, PGF2α and TXA2 in HUVEC. 10(-11)-10(-8)M DAA-I and 15min incubation specifically released PGE2 and PGI2. The release was inhibited by losartan and indomethacin but not by PD123319 and NS398 indicating that the angiotensin AT1 receptor and COX-1 mediate the release. At concentrations higher than 10(-7)M, DAA-I mimics the action of angiotensin II by releasing TXA2 but had no effect on the production of PGF2α. At similar concentrations and 4h incubation, DAA-I increased the release of the 4 prostaglandins via the angiotensin AT1 receptor and COX-2, again mimicking the action of angiotensin II. HUVEC that were preincubated with DAA-I or angiotensin II, released similar profiles of prostaglandins when incubated with arachidonic acid after the angiotensin had been washed off. We postulate that the internalized DAA-I/receptor complex remains active and mediates the conversion of arachidonic acid to the respective prostaglandins. The release of PGE2 and PGI2 via the angiotensin AT1 receptor and COX-1 is a novel specific action of DAA-I and is likely responsible for its beneficial effects seen in earlier studies. This specific action is definable as a biased agonism of the angiotensin AT1 receptor, which identifies DAA-I as a novel biased agonist and potential therapeutic that is able to produce specific prostaglandins at nanomolar concentrations.

Tryptase and protease-activated receptor-2 stimulate scratching behavior in a murine model of ovalbumin-induced atopic-like dermatitis.

The aim of the current study was to investigate the involvement of tryptase and protease-activated receptor-2 (PAR2) in the pathogenesis of itch using a recently developed murine model of atopic dermatitis (AD) elicited by epicutaneous sensitization with ovalbumin (OVA). We also examined whether tacrolimus exerts an antipruritic effect. Epicutaneous sensitization of BALB/c mice with OVA led to a significant increase in the number of scratches. Notably, PAR2 mRNA and protein levels as well as cutaneous levels of tryptase were significantly enhanced in epicutaneously sensitized mice. Pretreatment with the protease inhibitor, leupeptin, PAR2 antibody, and tacrolimus significantly reduced the number of degranulated mast cells and tryptase content, and consequently alleviated scratching behavior. Cetirizine (10mg/kg) exerted a significant inhibitory effect on the scratching behavior of mice, but did not affect the number of degranulated mast cells and induction of tryptase. Our results collectively suggest that tryptase and PAR2 are involved in OVA allergy-induced scratching behavior.

Cryptococcosis in kidney transplant recipients in a Chinese university hospital and a review of published cases.

BACKGROUND: Cryptococcosis is a severe fungal infection with a high mortality rate among solid-organ transplant recipients. Today, China is among the countries performing the most kidney transplants worldwide, however data on the association of cryptococcosis with kidney transplantation in mainland China remain scarce and fragmented. METHODS: We retrospectively analyzed cases of culture-confirmed cryptococcosis following kidney transplantation that have occurred at our hospital and reviewed the published cases in China over the last 30 years. RESULTS: Cryptococcosis in kidney transplant recipients was mainly caused by Cryptococcus neoformans var. grubii VNI strains and occurred most frequently in patients aged 41-50 years (37.9%, 11/29). The average time to infection after kidney transplantation was 5.16 ± 3.97 years. The clinical manifestations were found to be diverse, with slight to moderate headache and fever, meningeal irritation, and high cerebrospinal fluid pressure being relatively common. Physicians should be alert to these symptoms among kidney transplant recipients. CONCLUSIONS: Cryptococcosis is a serious infection among kidney transplant recipients in mainland China. It has unique characteristics, such as a relatively long time to onset after kidney transplantation, and diverse clinical manifestations. Treatment with intrathecal injection of amphotericin B is considered effective for central nervous system involvement. The findings of this study also highlight the urgent need for multicenter, prospective, and multidisciplinary clinical studies and education on cryptococcosis in kidney transplant recipients in China.

Des-aspartate-angiotensin I attenuates ICAM-1 formation in hydrogen peroxide-treated L6 skeletal muscle cells and soleus muscle of mice subjected to eccentric exercise.

L6 skeletal muscle cells overexpressed ICAM-1 when treated with H2O2. Maximum effect was observed at 200 µM H2O2. Des-aspartate-angiotensin I (DAA-I) concentration-dependently attenuated the overexpression. Maximum attenuation occurred at 10(-10) M DAA-I. H2O2 activated NFκB and its translocation into the nucleus of L6 muscle cells suggesting that NFκB mediates the H2O2-induced overexpression of ICAM-1. DAA-I inhibited the activation and translocation of NFκB. H2O2 is a major oxidant formed during skeletal muscle contraction and is implicated in oxidative stress and skeletal muscle damage in excessive unaccustomed exercise. The data show that DAA-I has antioxidant action, and its action was further investigated in the soleus muscle of mice subjected to 240 min of eccentric exercise on a rodent treadmill. The eccentric exercise induced superoxide formation and overexpression of ICAM-1 in the soleus muscle of the mice at 3 days post exercise. DAA-I (0.2 nmole/kg/day) administered orally on day 1 (pre-exercise) and 2 days post-exercise attenuated both the ROS formation and ICAM-1 overexpression. Earlier studies show that DAA-I acts as an agonist on the angiotensin AT1 receptor and elicits responses opposing those of angiotensin II. The present and earlier findings support the recent suggestion that angiotensin II is involved in skeletal muscle damage, and curtailment of its actions via ACE inhibitors and losartan protects and improves skeletal muscle damage. These findings open up new avenues for treatment and management of skeletal muscle damage via the interventions of the renin angiotensin system.

Comparison of clinical features in patients with persistent and nonpersistent cryptococcal meningitis: twelve years of clinical experience in four centers in China.

BACKGROUND AND AIMS: Cryptococcal meningitis (CM) has gradually increased in the recent 20 years in the whole world. Although the mortality decreased significantly in recent years, it was still high, especially in patients with persistent infection. Therefore, we compare differences of clinical features between persistent and nonpersistent CM patients. METHODS: We conducted a retrospective review of medical records of patients diagnosed with CM from January 2000 to December 2011 in four centers in China, including demographic features, underlying diseases, clinical presentations, laboratory data, and so on. RESULTS: Of 106 CM patients enrolled, 16 were identified as persistent cases. Among all variables, persistent CM patients were more like to be human immunodeficiency viruses (HIV) infection (P < 0.05), stiff neck (P < 0.01), a serum hemoglobin < 90 g/L (P < 0.01), a serum potassium concentration <2.7 mg/L (P < 0.01), an intracranial pressure (ICP) >400 mmH2 O (P < 0.01), and a latex agglutination cryptococcal antigen titer of cerebrospinal fluid (CSF LACT) >1:1024 (P < 0.01) than nonpersistent ones. A multivariate analysis showed that HIV infection (OR 7.49), stiff neck (OR 11.7), a serum potassium <2.7 mmol/L (OR 9.45), and an ICP >400 mmH2 O (OR 6.83) were closely correlated with persistent CM. CONCLUSIONS: Although it is difficult to deal with persistent CM nowadays, some cases could be predicted early enough in the future, so as to be treated appropriately and have relatively good outcomes.

Association of vitamin D receptor FokI and ApaI polymorphisms with human cytomegalovirus disease in the first three months following kidney transplantation.

BACKGROUND: Untreated human cytomegalovirus (CMV) disease (CMVD) is an identified risk factor for reduced rates of patient (and graft) survival, death or retransplantation in kidney transplant recipients due to increased immunological tolerance after transplant. Vitamin D receptor (VDR) gene polymorphisms have an obvious relationship with autoimmune diseases but the relationship between VDR gene polymorphisms and CMVD are not well understood. This study investigated the relationship between VDR FokI and ApaI gene polymorphisms and CMVD, and their value for predicting risk of CMVD. METHODS: Ninety-eight kidney transplantation recipients were randomly chosen for which peripheral blood samples and case histories for the first three months after kidney transplantation were obtained. Using polymerase chain reaction-restriction fragment length polymorphisms, 30 recipients were found to be homozygous for the FokI gene (FF), 47 heterozygous (Ff), and 21 were homozygous (ff). Likewise, similar analyses determined that 12 recipients were homozygous for the ApaI gene (AA), 36 heterozygous (Aa), and 50 homozygous (aa). Factors affecting the prognosis of the kidney transplantation were compared for all genotypes by statistical analysis before operation. Infection by CMV for all recipients was detected by immunofluorescence assay to diagnose CMVD. RESULTS: No statistical significance was observed for the factors affecting the prognosis of the kidney transplantation between both genotypes; however, statistical differences in CMVD among the FokI genotypes were identified. It was determined that the risk of CMVD was significantly increased for recipients of the ff genotype than for other genotypes. There was no statistical significance observed for CMVD among ApaI genotypes. CONCLUSIONS: The recessive f allelic gene of VDR can be regarded as a risk factor of CMVD while FF recipients have lower incidence of CMVD after kidney transplantation. ApaI genotypes showed no relationship with predisposition to CMVD.

Disseminated cryptococcal lymphadenitis with negative latex agglutination test.

We reported an unusual case of disseminated cryptococcal lymphadenitis in an immunocompetent host who presented with fever and lymphadenopathy, which were the only two symptoms and signs. Latex agglutination test of serum and cerebrospinal fluid (CSF) were negative, while lymph node biopsy showed Cryptococcus neoformans. A diagnosis of disseminated cryptococcal lymphadenitis was made. Then the patient was treated with amphotericin B for 15 days as initial therapy and itraconazole for 6 months as maintenance therapy respectively. The patient received re-examination per 6 months and was followed up for 2 years. Swollen lymph nodes diminished gradually, and no fever or other symptoms were found. Latex agglutination test of serum and CSF were negative throughout the follow-up period, and anti-HIV, syphilis and tuberculosis antibody were all negative.

Xiaoyin recipe ( ) for psoriasis induces a Th1/Th2 balance drift toward Th2 in peripheral blood mononuclear cells of experimental autoimmune thyroiditis rats.

OBJECTIVE: To explore the regulatory mechanism of Xiaoyin Recipe () on the T helper 1/T helper 2 (Th1/Th2) immune balance. METHODS: Thirty-six experimental animals were divided into three groups, 12 rats in each group: blank control group (B group), negative control group (N group), and Xiaoyin Recipe treatment group (T group). The latter two groups received immunization of experimental autoimmune thyroiditis (EAT), and T group were treated with Xiaoyin Recipe for a month. Then, the expression of Th1-Th2-related genes in peripheral blood mononuclear cells (PBMCs) were screened with Oligo GEArray Rat Th1-Th2-Th3 Microarray. The expressions of tumor necrosis factor-α (TNF-α), interleukin-10 (IL-10), T-box expressed in T-cells (T-bet), and GATA-binding protein-3 (GATA-3) were detected by real-time polymerase chain reaction (RT-PCR). RESULTS: Gene array screening showed that compared to N group, in T group after Xiaoyin Recipe treatment, 3 genes were upregulated in EAT rats, including interleukin-27 receptor alpha (IL-27rα), glomulin (Glmn), and GATA-3, while 38 genes were downregulated, such as CD28, IL-18, signal transducer, and activator of transcription 1 (STAT1), T-bet, TNF receptor superfamily member 4 (TNFRSF4), TNF ligand superfamily member 5 (TNFSF5), and TNF receptor superfamily member 5 (TNFRSF5). While RT-PCR showed that there was an increased level of TNF-α mRNA (P<0.01), an elevated ratio of T-bet/GATA-3, and a decreased level of IL-10 mRNA in PBMC of N and T group compared to B group (P <0.01); and after treatment with Xiaoyin Recipe, IL-10 mRNA level increased (P <0.01), while TNF-α mRNA level and T-bet/GATA-3 ratio decreased in T group compared to N group (P <0.01). CONCLUSION: Xiaoyin Recipe for psoriasis could induce a Th1/Th2 balance drift toward Th2 in PBMC of EAT rats and thus improve the conditions.

[Up-regulation of the sHLA-G5 level by human-mouse chimeric anti-IL-2R monoclonal antibody treatment in kidney transplantation].

OBJECTIVE: To find whether the up-regulation of soluble human leucocyte antigen-G5 (sHLA-G5) levels is a new function mechanism of anti-interleukin-2 receptors (anti-IL-2R) monoclonal antibody treatment in kidney transplantation. METHODS: A total of 215 recipients at our centre from January 2006 to December 2007 were divided into antibody use group (n = 141) and antibody non-use group (n = 74) and another healthy group (n = 69). The sHLA-G5 level in peripheral blood was detected by enzyme-linked immunosorbent assay (ELISA). And the expression of HLA-G5 was confirmed by Western blot and Real-time polymerase chain reaction (PCR). RESULTS: sHLA-G5 levels was (56 ± 30) µg/L in using anti-IL-2 receptor monoclonal antibody before transplantation, It was higher than that before use antibody [(34 ± 20) µg/L], also higher than healthy group [(35 ± 17) µg/L] and antibody non-use group [(36 ± 19) µg/L, P < 0.05, respectively]. At Day 1, Day 4, Week 1, Week 2 post-transplantation, the level of sHLA-G5 of recipients with antibody use was significantly higher than that of those with antibody non-use. The values were as follows: (95 ± 35) µg/L vs (54 ± 16) µg/L, (131 ± 24) µg/L vs (75 ± 22) µg/L, (167 ± 44) µg/L vs (62 ± 17) µg/L, (172 ± 35) µg/L vs (45 ± 16) µg/L (all P < 0.01). And the results of Western blot and RT-PCR corresponded to those of ELISA. CONCLUSION: The preoperative use of first dose of anti-IL-2R monoclonal antibodies results in the up-regulated level of sHLA-G5. Thus it is beneficial for protecting the kidney survival and reducing the risks of acute rejection.

[Identification and functional study of Tim-1(+)CD19(+) regulatory B cell in kidney transplantation recipients].

OBJECTIVE: To observe the ratio of Tim-1(+)CD19(+) B cell in the peripheral blood of kidney transplantation recipients and elucidate its functions. METHODS: From December 2009 to June 2010, a total of 35 pairs of kidney transplant recipients were selected and divided into 3 groups: healthy donors as control (n = 35), pre-transplantation (n = 35) and post-transplantation (n = 35). The profiles of Tim-1(+)CD19(+) B cell in kidney transplantation donors and recipients were analyzed and sorted by flow cytometry (FCM). Mixed lymphocyte culture (MLC) was carried out between kidney transplantation donors and recipients. After the additions of Tim-1(+)CD19(+) and Tim-1(-)CD19(+) B cells, there were 3 groups: Tim-1(+), Tim-1(-) and blank. Lymphocyte proliferation and inhibition status were evaluated by propidium iodide uptake and Annexin V binding. And the cytokine levels were detected by FCM. RESULTS: The absolute values of peripheral CD19(+)B cells were (170 ± 90), (202 ± 99), (155 ± 71) cells/µl in the pre-transplantation, post-transplantation and control groups respectively, post-transplantation group were higher than control group (P = 0.0300). The Tim-1(+)CD19(+) cell ratios were (2.20 ± 0.98)%, (35.46 ± 10.66)% and (1.95 ± 0.95)% in three groups. And the differences were statistically significant (both P < 0.01). Tim-1(+)CD19(+) B and Tim-1(-)CD19(+) B cells were added into MLC respectively. The early apoptotic cells of the Tim-1(+) group were higher than those in the Tim-1(-) group [(45.31 ± 12.37)% vs (10.92 ± 2.14)%, P < 0.05] and significantly higher than the blank group [(1.93 ± 0.26)%, P < 0.01]. Late apoptotic and dead cells of the Tim-1(+) group were higher than those in the Tim-1(-) group [(21.32 ± 5.67)% vs (2.32 ± 0.31)%, P < 0.01] and the blank group [(1.27 ± 0.19)%, P < 0.05]. The interleukin 10 levels in MLC supernatant of the Tim-1(+) group were significantly higher than those in the Tim-1(-) group [(5.32 ± 0.37) pg/ml vs (2.46 ± 0.25) pg/ml, P = 0.0001]. However, the interferon-γ levels were lower than those in the Tim-1(-) group [(1.51 ± 0.22) pg/ml vs (4.69 ± 0.32) pg/ml, P = 0.0015]. CONCLUSION: Present in the peripheral blood of kidney transplantation recipients, Tim-1(+)CD19(+) B cell has the capacity of promoting lymphocytic apoptosis. As a new regulatory subset of B cells, it plays important roles in the immune responses of transplantation.

[Applications of monitoring human leukocyte antigen G and its inhibitory receptors in post-renal transplantation].

OBJECTIVE: to study the feasibility of human leucocyte antigen-G (HLA-G) as a post-transplantation prognostic biomarker and discuss the correlation of its receptor expression and the mechanisms. METHODS: a total of 215 recipients in our centre from February 2006 to June 2008 were divided into stable kidney function group (n = 173) and acute rejection group (n = 42). The soluble human leucocyte antigen-G5 (sHLA-G5) level in peripheral plasma was detected by ELISA. And the HLA-G receptor ILT-2, KIR2DL4 on T, B, NK lymphocytes were analyzed by flow cytometry (FCM). The sHLA-G5 cutoff level by ROC curve was employed to predict the events of acute post-transplantation rejection. And regression analysis was used to determine the association of sHLA-G5 with acute rejection. RESULTS: an optimal cutoff value of 139.0 microg/L could be defined for sHLA-G5 (sensitivity: 63.6%, specificity: 82.1%, AUC: 0.780). Binary regression analysis showed that sHLA-G5 played an independent role on acute rejection (P = 0.019, OR = 0.039, 95%CI: 2.091 - 5.661). The rate of HLA-G receptor ILT-2 on CD4(+)T cell, CD8(+)T cell and B cell in acute rejection group was statistically lower than that in stable kidney function group (21% ± 7% vs 52% ± 17%, 23% ± 6% vs 39% ± 16%, 21% ± 7% vs 39% ± 16%, all P < 0.05). The expression of KIR2DL4 on NK cells in acute rejection group was statistically lower than that in stable kidney function group (31% ± 10%vs 57% ± 21%, P < 0.05). CONCLUSION: sHLA-G5 level may be predicted for acute rejection with a high sensitivity and specificity. The up-regulated expression of ILT-2 and KIR2DLT may contribute to immunology tolerance in peripheral circulation.

[The application of quantitative detection of cystatin C in evaluation of renal function after renal transplantation].

AIM: To establish ELISA method for quantitate the concentration of cystatin C (cys C) and to monitor the renal function of patients before and after renal transplantation. METHODS: Hybridomas secreting monoclonal antibodies (mAbs) against human cys C were produced and sandwich ELISA kit for quantitatively detecting cys C was established. Then the concentrations of serum cystatin C (Scys C) and urine cystatin C (Ucys C) from normal controls and 23 patients undergoing renal transplantation were detected and their relationship with serum creatinine (SCR) was analyzed. RESULTS: Seven hybridomas secreting anti-cys C mAbs were obtained. The sensitivity of the established ELISA kit reached 0.1 µg/L. The concentrations of Scys C and Ucys C of normal healthy controls were in accordance with other report. High correlations between Scys C or Ucys C and the level of SCR were observed (P<0.01). Rapid decline of Scys C and Ucys C concentrations was consistent with the decrease of SCR in the patients with normal course (NC) recovery after renal transplantation. However, Ucys C kept higher level within two weeks after the operation in patients with AR until the day 21. In patients with DGF, higher levels of Scys C, Ucys C and SCR were sustained within four weeks after renal transplantation. CONCLUSION: The sensitive ELISA kit for detection of cys C has been established. Importantly, there are the persistently high levels of Scys C and Ucys C in patients with AR or DGF, which can be used as a novel indicator for monitoring renal function after renal transplantation.

[Expression of human leucocyte antigen G in peripheral blood of kidney transplantation recipients].

OBJECTIVE: To investigate the expression of non-classical major histocompatibility complex (MHC)-I molecule, human leucocyte antigen (HLA) G, including membrane-bound HLA-G (mHLA-G), intracellular HLA-G (iHLA-G) and soluble HLA-G (sHLA-G), in peripheral blood of surviving kidney transplantation recipients and understand the relevance between HLA-G and the function of transplanted organ, as well as the onset of acute rejection. METHODS: A longitudinal study was performed on 175 kidney transplantation recipients. Three groups were involved in this study, including acute rejection group (n = 36), function stable group (n = 139) and healthy control group (n = 30). The expression of mHLA-G1 and iHLA-G1 in the T lymphocytes of peripheral blood was detected by flow cytometry analysis and the sHLA-G5 level detected by ELISA. RESULTS: The average rate of CD4(+)mHLA-G1(+), CD8(+)mHLA-G1(+), CD4(+)iHLA-G1(+), CD8(+)iHLA-G1(+) in T lymphocytes of healthy control group was 0.43% +/- 0.19%, 1.23% +/- 0.41%, 27% +/- 13% and 36% +/- 14% respectively. That of acute rejection group was 0.57% +/- 0.34%, 1.31% +/- 0.56%, 26% +/- 8% and 37% +/- 17%; that of function stable group was 0.61% +/- 0.43%, 1.39% +/- 0.47%, 26% +/- 9% and 37% +/- 17% respectively. There was no significant difference among the three groups (all P > 0.05). The average of sHLA-G5 levels in plasma of control group was (25 +/- 14) ng/ml, acute rejection group (24 +/- 15) ng/ml (pre-operative) and (34 +/- 21) ng/ml (post-operative), function stable group (25 +/- 11) ng/ml (pre-operative) and (56 +/- 32) ng/ml (post-operative). There was no significant difference among the three groups (pre-operative, P > 0.05). The average of sHLA-G5 levels in plasma of function stable group was higher than that of acute rejection group (post-operative, P < 0.05). CONCLUSION: There is a subset of CD4(+)HLA-G1(+) and CD8(+)HLA-G1(+)T lymphocytes with low percentage in peripheral blood of those surviving kidney transplantation recipients. The expressions of mHLA-G1 and iHLA-G1 have no relevance with the onset of acute rejection. sHLA-G5 is correlated with acute rejection in peripheral blood of surviving transplantation recipients.