Description of three new Boulenophrys species from eastern Guangdong, China, emphasizing the urgency of ecological conservation in this region (Anura, Megophryidae).

The hilly region in eastern Guangdong, China lacks comprehensive scientific investigations for decades, especially in terms of herpetofauna. In recent years, several highly threatened amphibians have been gradually discovered from this region. In this work, three new species of the genus Boulenophrys are described, which are endemic from only one or two known localities in eastern Guangdong. These discoveries enrich the diversity of Boulenophrys in eastern Guangdong. With the large number of threatened urodeles and anuran species occurring in this densely populated area, the unique herpetological diversity in eastern Guangdong is facing the impacts of habitat degradation and fragmentation, and conservation actions are urgently required.

A milbemycin compound isolated from Streptomyces Sp. FJS31-2 with cytotoxicity and reversal of cisplatin resistance activity in A549/DDP cells.

Streptomyces Sp FJS31-2 is a strain isolated from special habitat soils in the early stage of our laboratory for producing a new type of halogenated type II polyketide antibiotic with good anti-MRSA activity. In this experiment, a variety of chromatographic and spectroscopic methods was used to isolate and identify a milbemycin compound VM48130 from the ethyl acetate extract of the fermentation products. To investigate its bioactivity, Cell Counting Kit-8 (CCK-8) assay was used to test the cytotoxic activity of the compound against a variety of cancer cells (human liver cancer cell line MHCC97H and SK-Hep1, human nasopharyngeal carcinoma cell line CNE1, mouse melanoma cell line B16, human colon cancer cell line LOVO, human lung adenocarcinoma cell line A549) and normal cells (human bronchial epithelial cell line 16HBE, human normal liver cell line L02, human nasopharyngeal epithelial cell line NP69). The results showed that the compound had significant cytotoxic activity against the above cancer cells, and the IC50 values were 21.96 ± 1.45, 22.18 ± 0.55, 19.42 ± 0.71, 18.61 ± 1.68, 18.62 ± 0.67, 18.52 ± 0.64 µM, respectively. Furthermore, the CCK-8 method was used to evaluate the compound's reversal of cisplatin resistance in multidrug resistant cisplatin-resistant human lung adenocarcinoma (A549/DDP) cells. The results indicated that when the compound concentration was 0.5 µM, the reversal fold (RF) reached 6.25 and showed a dose-dependent effect. At 5 µM, the RF reached 8.35, which was approximately equivalent to the reversal effect of the positive drug verapamil at the same concentration. The expression of MDR1, MRP1, LRP, MAST1 resistance genes and the corresponding proteins were analyzed by quantitative RT-PCR and Western blot assay, and found that the compound could significantly down-regulate the expression of these genes and proteins. These results indicated that VM48130 had the potential of being a lead compound for the treatment or adjuvant treatment of cancer.

[Overexpression of eIF4E gene in acute myeloid leukemia and its relation with disease progression].

The aim of this study was to detect the expression level of eIF4E gene in patients with non-treated, remission and non-remission/relapse acute myeloid leukemia (AML), and other non-malignant haematologic diseases so as to analyze and reveal the relationship of eIF4E gene expression with AML progression. SYBR Green I RT-PCR was used to assay the expression level of eIF4E mRNA extracted from bone marrow mononuclear cells in 30 patients with AML (6 in M2, 5 in M3, 8 in M4, 10 in M5, 1 in M6) and 20 patients with non-malignant hematologic diseases. The ß2-microglubin(ß2M) was used as internal reference and the formula 2(-ΔCt)×100% was applied to calculate the expression level of eIF4E gene. The results showed that the eIF4E expression level (7.098 ± 5.544)% in patients with non-treated and non-remitted/relapsed AML was significantly higher than that in patients with remission (0.964 ± 0.312)% (P < 0.01) and non-malignant hematologic diseases (0.248 ± 0.163)% (P < 0.01). There was no difference between latter two group patients, even though the expression level of eIF4E gene in patients with M4 and M5 was higher. As compared with non-malignant hematologic diseases, the expression level of eIF4E gene of patients with remission patients showed no significant difference. It is concluded that the over-expression of eIF4E gene has been found in patients with AML, and its level obviously decreases along with remission of disease, thus the eIF4E gene may be a surveillance parameter for disease progression.

Jiangxienone, a new compound with potent cytotoxicity against tumor cells from traditional Chinese medicinal mushroom Cordyceps jiangxiensis.

A new compound, named jiangxienone, has been isolated from a culture of the traditional Chinese medicinal mushroom Cordyceps jiangxiensis, and its chemical structure was established on the basis of spectroscopic and chemical techniques. Jiangxienone showed potent cytotoxic effects against human gastric adenocarcinoma SGC-7901 cell and human lung carcinoma A549 cell with IC(50) values ranging from 1.38 to 2.93 µM, i.e., with at least approximately six-fold stronger cytotoxicity than cisplatin, a first-line chemotherapy drug for cancer patients.

[Analysis of TCR gene rearrangement for identification of T cell leukemia clone by using fine-tiling aCGH].

AIM: To establish a new method which analyzes T cell receptor (TCR) gene rearrangement for identification of T cells acute lymphoblastic leukemia (T-ALL) clone, it will provide the basis for the study of T-ALL including the chromosome translocation involving TCR loci. METHODS: Total DNA was isolated from peripheral blood mononuclear cells (PBMC) of one case with T-ALL. Using the fine-tiling array comparative genomic hybridization (fine-tiling aCGH) to analyze the genomic DNA differences of the case and control group, we could find the breakpoints and their position in the chromosomes. According to the preliminary results, we could design the specific primers for the positions of the breakpoints relative to sequence. Furthermore, the ligation-mediated PCR (LM-PCR) and sequence analysis were used to identify the TCR gene rearrangement. And TCR gene expression was detected by RT-PCR. RESULTS: The fine-tiling aCGH results of the T-ALL showed that the TCRα/δ locus of chromosome 14 appeared four breakpoints, corresponding to TCR Vδ1, Vδ2, Jδ1 and Jδ2. By LM-PCR, sequencing and sequence analysis, TCR gene of the case of T-ALL was involved in Vδ1Dδ2Dδ3Jδ1, Vδ2Dδ3Jδ2 rearrangement. RT-PCR results also confirmed the expression of these TCR gene rearrangements. CONCLUSION: The results demonstrated that fine-tiling aCGH and LM-PCR techniques could be used to identify the TCR gene rearrangement as one of the best perfect methods. And it was also a way to find some fusion genes involving in TCR gene.

Mechanism of multidrug resistance of human small cell lung cancer cell line H446/VP.

BACKGROUND: Small cell lung cancer (SCLC) is the most aggressive form of lung cancer. This study aimed to investigate the mechanism of human small cell lung cancer cell line resistance to etoposide (VP-16), H446/VP. METHODS: The cell viability was measured by MTT assay. Immunocytochemistry, reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting methods were used to detect the multidrug resistance gene (MDR1), bcl-2, bax and the topoisomerase II (Topo II) expressions in H446 and H446/VP cells after treated with or without VP-16. RESULTS: The 50% inhibition concentration (IC50) of VP-16 on H446 cells was 49 mg/L, and 836 mg/L was for H446/VP cells. The expressions of MDR1 and bcl-2 were up-regulated, while the amounts of bax and Topo II were reduced in H446/VP cells. After treated with 49 mg/L of VP-16, it showed that the drug could significantly inhibit bcl-2 and Topo II expressions, and increase bax expression in H446 cells compared with that of H446/VP cells. CONCLUSIONS: The H446/VP cell was stably resistant to VP-16. The decreased expression of Topo II was correlated with the H446/VP multidrug resistance. The elevated expressions of MDR1, and the altered apoptotic pathways also played an important role in VP-16 induced multidrug resistance of SCLC.

[Change of expression pattern of CD3 genes in peripheral blood T-cells from CML patients].

Our previous finding showed that down-regulation of CD3ζ gene was detected in patients with chronic myeloid leukemia (CML). In order to further elucidate the feature of T cell immune status in the signal transduction in CML patients, the expression patterns of all 4 CD3 genes were characterized in peripheral blood of patients, the expression levels of CD3γ, δ, ε and ζ chain genes were detected by real time qPCR with SYBR Green I staining in peripheral blood mononuclear cells (PBMNCs) from 17 cases of de novo CML patients in chronic phase and 17 cases of healthy individuals, the ß2-microglobulin gene was used as an internal reference, and the mRNA expression level of each CD3 gene was evaluated by the 2(-ΔCt) x 100% method. The results showed that the median expression levels of CD3γ, δ and ε genes (2.344%, 0.515% and 3.516%) in CML patients were not significantly different from healthy individuals (p = 0.072, p = 0.190, p = 0.615, respectively), while the expression level of CD3ζ gene in PBMNCs from CML patients (0.395%) was lower than that from healthy individuals (1.538%) (p < 0.001). The expression patterns of 4 CD3 genes in proper order were CD3ε > CD3γ > CD3δ > CD3ζ in CML group, in contrast, the expression patterns were presented as CD3γ > CD3ε > CD3ζ > CD3δ in healthy group. It is concluded that the present study characterized the expression pattern of CD3γ, δ, ε and ζ chain genes in CML patients, lower expression of CD3ζ is the feature of TCR signal transduction immunodeficiency and the expression patterns of 4 CD3 genes are changed in CML patients.

Incidence, risks, and outcome of idiopathic pneumonia syndrome early after allogeneic hematopoietic stem cell transplantation.

We retrospectively analyzed 23 cases with early-onset idiopathic pneumonia syndrome (IPS) of 192 patients undergoing allogeneic hematopoietic stem cell transplantation (HSCT) from April 1997 to October 2007. Risk factors for IPS development were evaluated using Cox proportional hazards model, including age, gender, underlying disease, disease status at transplant, transplant type, conditioning regimens, donor type, acute graft-vs.-host disease (GVHD), severity of acute GVHD (aGVHD), human leukocyte antigen (HLA) disparity, and organ involvement of aGVHD. Factors that were significant at the 0.1 level on univariate analysis were evaluated by multivariate analysis. Twenty-three of 192 patients developed IPS (12.0%). Median time to IPS onset after allogeneic HSCT was 76 d (range 32-120 d); median time to death after the diagnosis of IPS was 9 d (range 3-92 d); 20 patients with IPS died because of the rapid progression of respiratory failure (87.0%). Nineteen patients with IPS developed aGVHD (82.6%), with grade III-IV aGVHD in 11 patients (47.8%) and aGVHD of gut in 16 patients (69.6%). The following six factors were associated with an increased risk of IPS by univariate analysis: not in remission, unrelated donor, HLA disparity, occurrence of aGVHD, grade III-IV aGVHD and aGVHD of gut. These risk factors were entered into a multivariate analysis model. Only unrelated donor, grade III-IV aGVHD and aGVHD of gut are identified as being significantly associated with the occurrence of IPS, and among them, aGVHD of gut was associated with the largest risk of IPS, suggesting that the lung may be a target organ of aGVHD.

[Detection of TCR Vbeta subfamily sjTRECs in normal peripheral blood and cord blood].

AIM: To detect the existence of sjTRECs in TCR 23 Vbeta subfamilies in normal peripheral blood and cord blood, and to evaluate the recent thymic emigrants of naive T cells of different TCRbeta subfamilies. METHODS: Different amounts of DNA from samples (4 cases of thymocytes, 10 cases of cord blood and 10 cases of normal PBMCs) were amplified to estimate the frequency of 23 TCR Vbeta-Dbeta sjTRECs by using semi-nest PCR. RESULTS: At the same cellular concentration, the most frequency of Vbeta-Dbeta1 sjTRECs was found in thymocytes, the second was in cord blood, and the lowest was in peripheral blood. At 2x10(5), 5x10(4) and 1x10(4) cells level, sjTRECs of most Vbeta subfamilies could be detected in cord blood and peripheral blood, while sjTRECs of some Vbeta subfamilies could be detected at 1x10(3) cells level. The frequencies of Vbeta2-, Vbeta4-, Vbeta7-, Vbeta11- and Vbeta19-Dbeta1 sjTRECs and the number of detectable Vbeta subfamilies from normal PBMCs were obviously lower than those from cord blood. CONCLUSION: The technique of semi-quantitive PCR to detect sjTRECs in 23 Vbeta subfamilies was successfully established. Naive T cells from different Vbeta subfamilies could be detected with different frequency.

Management of star fruit-induced neurotoxicity and seizures in a patient with chronic renal failure.

An 84-year-old Asian woman with hypertension and chronic renal failure was evaluated for incoherent speech, followed by intermittent interruptions of consciousness, and then status epilepticus after ingesting one star fruit (Averrhoa carambola) each day for 3 days. Conventional first-line anticonvulsants and hemodialysis were administered without significant control of the patient's seizures. Treatment was started with propofol, an intravenous agent that induces anesthesia with rapid onset and elimination from the central nervous system; this resulted in complete control of the seizures. Propofol may be an effective alternative when dialysis and conventional first-line anticonvulsants are unsuccessful in treating the symptoms of neurotoxicity.

[Analysis of selected usage and clonal expansion of TCR Vbeta repertoire of peripheral blood T cells in patients with non-Hodgkin's lymphoma].

To investigate the distribution and clonal expansion of TCR Vbeta subfamily T cells in patients with B-NHL and T-NHL, the CDR3 of TCR Vbeta 24 subfamily genes was amplified in peripheral blood mononuclear cells from 4 cases with B-NHL and 2 cases with T-NHL using RT-PCR, and to observe the usage of TCR Vbeta repertoire, the PCR products were further labeled with fluorescence and analyzed by genescan technique for the CDR3 size, to evaluating clonality of the detectable TCR Vbeta T cells. The results indicated that only selected expression of 6-12 Vbeta subfamily T cells could be identified in the 6 cases with NHL, and Vbeta1, Vbeta8, Vbeta13 and Vbeta19 were expressed in all samples, Vbeta2 and Vbeta16 could be found in 5 samples, whereas Vbeta4-6, Vbeta10-12, Vbeta15, Vbeta17-18, Vbeta20 and Vbeta22-23 were absent in all samples. Genescan analysis showed that clonal expansion of T cells could be found in 1-3 Vbeta subfamilies from 2 cases with B-NHL and 1 case with T-NHL. In conclusions, the similar selected usage of TCR Vbeta subfamily T cells could be found in peripheral blood from patients with B and T NHL, clonal expansion of T cells which were considered to be related to lymphoma cell antigen could be detected in a part of patients with B or T NHL.

[The feature of TCR V alpha40 with Jdelta1, Ddelta3 or psiJalpha gene rearrrangements in T cells].

The rearrangement segments of TCR Valpha40 gene with Jdelta1, Ddelta3 or psi Jalpha were amplified in genomic DNA from peripheral blood mononuclear cells of 10 normal subjects, sorted CD3(+) cells from peripheral blood of 4 cases and thymocytes from 7 cases, by using nested PCR. Different amounts of DNA from all samples were amplified to estimate the frequency of Valpha40 gene rearrangements. The results indicated that the rearrangements of TCR Valpha40 gene with Jdelta1, Ddelta3 or psi Jalpha could be found respectively in the most samples of peripheral blood T cells or thymocytes. The frequencies of Valpha40 rearrangements were different in peripheral blood T cells and thymocytes by analysis of PCR with different amounts DNA. It is concluded that the TCR V alpha40-psi Jalpha was the most frequent rearrangement in mature and immature T cells, whereas TCR Valpha40-Ddelta3 was more frequently rearranged in immature T cells