Ameliorative effect of anisodamine (654-1/654-2) against myocardial dysfunction induced by septic shock via the NF-κB/NLRP-3 or the PI3K-AKT/NF-κB pathway.

BACKGROUND: Septic shock, an extremely dangerous condition that causes impairment of organ function, always largely contributes to mortality in intensive care units. The impact of septic shock-induced organ damage on morbidity and mortality is substantially influenced by myocardial dysfunction. However, it remains unclear whether and in what manner anisodamine (654-1/654-2) ameliorates myocardial dysfunction caused by septic shock. PURPOSE: This study is the pioneering investigation and validation about the protective efficacy of anisodamine (654-1/654-2) against LPS-induced myocardial dysfunction in septic shock rats. It also aims to explore the differences in the underlying molecular mechanisms of both drugs. METHODS: A septic shock model was established in SD rats by after tail vein administration of LPS. 64 rats were distributed into eight groups, such as LPS group, control group, LPS+654-1 group (1.25, 2.5, and 5 mg/kg), and LPS+654-2 group (1.25, 2.5, and 5 mg/kg). The hemodynamics, echocardiography, immunohistochemical analysis, TEM, TUNEL assay, and H&E staining were utilized to assess the septic shock model and myocardial function. Lactic acid, inflammatory markers (IL-1ß, IL-6, and TNF-α), endothelial injure markers (SDC-1, HS and TM) and myocardial injury markers (CK, c-TNT and NT-pro BNP) were assessed using ELISA or biochemical kits. Additionally, the mechanisms of 654-1/654-2 were analyzed using RNA-seq and bioinformatics, and validated using western blotting and RT-PCR. RESULTS: Administration of 654-1/654-2 significantly restored hemodynamics and improved myocardial and endothelial glycocalyx injury in septic shock rats. Furthermore, 654-1/654-2 dose-dependently reduced plasma levels of lactic acid, inflammatory cytokines, and markers of endothelial and myocardial injury. Analyses using RNA-seq, WB and RT-PCR techniques indicated that 654-1/654-2 could mitigate myocardial and endothelial injury by inhibiting the NF-κB and NLRP-3 pathways, and activating the PI3K-AKT pathway. CONCLUSIONS: These findings demonstrated that 654-1/654-2 could alleviate myocardial damage in septic shock rats. Specifically, 654-1 inhibited the NF-κB/NLRP-3 pathway, whereas 654-2 promoted the PI3K-AKT pathway and inhibited the NF-κB pathway, effectively mitigating the inflammatory response and cell apoptosis.

Targeting endothelial cells with golden spice curcumin: A promising therapy for cardiometabolic multimorbidity.

Cardiometabolic multimorbidity (CMM) is an increasingly significant global public health concern. It encompasses the coexistence of multiple cardiometabolic diseases, including hypertension, stroke, heart disease, atherosclerosis, and T2DM. A crucial component to the development of CMM is the disruption of endothelial homeostasis. Therefore, therapies targeting endothelial cells through multi-targeted and multi-pathway approaches hold promise for preventing and treatment of CMM. Curcumin, a widely used dietary supplement derived from the golden spice Carcuma longa, has demonstrated remarkable potential in treatment of CMM through its interaction with endothelial cells. Numerous studies have identified various molecular targets of curcumin (such as NF-κB/PI3K/AKT, MAPK/NF-κB/IL-1ß, HO-1, NOs, VEGF, ICAM-1 and ROS). These findings highlight the efficacy of curcumin as a therapeutic agent against CMM through the regulation of endothelial function. It is worth noting that there is a close relationship between the progression of CMM and endothelial damage, characterized by oxidative stress, inflammation, abnormal NO bioavailability and cell adhesion. This paper provides a comprehensive review of curcumin, including its availability, pharmacokinetics, pharmaceutics, and therapeutic application in treatment of CMM, as well as the challenges and future prospects for its clinical translation. In summary, curcumin shows promise as a potential treatment option for CMM, particularly due to its ability to target endothelial cells. It represents a novel and natural lead compound that may offer significant therapeutic benefits in the management of CMM.

Alpinia katsumadai Hayata Volatile Oil Is Effective in Treating 5-Fluorouracil-Induced Mucositis by Regulating Gut Microbiota and Modulating the GC/GR Pathway and the mPGES-1/PGE2/EP4 Pathways.

This study was aimed to investigate the therapeutic effect and mechanism of AKHO on 5-fluorouracil (5-FU)-induced intestinal mucositis in mice. Mouse body weight, diarrhea score, and H&E staining were applied to judge the therapeutic effect of AKHO. 16S rDNA and nontargeted metabolomics have been used to study the mechanism. WB, ELISA, and immunohistochemistry were adopted to validate possible mechanisms. The results demonstrated that AKHO significantly reduced diarrhea scores and intestinal damage induced by 5-FU in mice. AKHO lowered the serum levels of LD and DAO, and upregulated the expressions of ZO-1 and occludin in the ileum. Also, AKHO upregulated the abundance of Lactobacillus in the gut and suppressed KEGG pathways such as cortisol synthesis and secretion and arachidonic acid metabolism. Further validation studies indicated that AKHO downregulated the expressions of prostaglandin E2 (PGE2), microsomal prostaglandin E synthase-1 (mPGES-1), and PGE2 receptor EP4, as well as upregulated the expression of glucocorticoid (GC) receptor (GR), leading to improved intestinal epithelial barrier function. Taken together, AKHO elicited protective effects against 5-FU-induced mucositis by regulating the expressions of tight junction proteins via modulation of GC/GR and mPGES-1/PGE2/EP4 pathway, providing novel insights into the utilization and development of this pharmaceutical/food resource.

Scutellarin suppresses triple-negative breast cancer metastasis by inhibiting TNFα-induced vascular endothelial barrier breakdown.

Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer with high vascularity and frequent metastasis. Tumor-associated abnormal vasculature was reported to accelerate TNBC metastasis. Scutellarin (SC) is a natural flavonoid with a cardiovascular protective function. In this study, SC reduced TNBC metastasis and alleviated tumor-associated vascular endothelial barrier injury in vivo. SC rescued the tumor necrosis factor-α (TNFα)-induced diminishment of endothelial junctional proteins and dysfunction of the endothelial barrier in vitro. SC reduced the increased transendothelial migration of TNBC cells through a monolayer composed of TNFα-stimulated human mammary microvascular endothelial cells (HMMECs) or human umbilical vein endothelial cells (HUVECs). TNFα induced the nuclear translocation of enhancer of zeste homolog-2 (EZH2), and its chemical inhibitor GSK126 blocked TNFα-induced endothelial barrier disruption and subsequent TNBC transendothelial migration. TNF receptor 2 (TNFR2) is the main receptor by which TNFα regulates endothelial barrier breakdown. Extracellular signal-regulated protein kinase (ERK)1/2 was found to be downstream of TNFα/TNFR2 and upstream of EZH2. Additionally, SC abrogated the TNFR2-ERK1/2-EZH2 signaling axis both in vivo and in vitro. Our results suggest that SC reduced TNBC metastasis by suppressing TNFα-initiated vascular endothelial barrier breakdown through rescuing the reduced expression of junctional proteins by regulating the TNFR2-ERK1/2-EZH2 signaling pathway.

Do foreign direct investment and environmental regulation improve green technology innovation? An empirical analysis based on panel data from the Chinese manufacturing industry.

The environmental regulation and foreign direct investment (FDI) inflow have an important impact on the progress of green technology. This study analyzes the impacts of environmental regulation and FDI on green technology innovation (GTI) based on the panel data of 13 Chinese manufacturing sectors. The results of static panel regression show that the environmental regulation has a positive impact on GTI, while the FDI has a negative impact. The results of the panel threshold model reveal that the effect of environmental regulation on GTI presents a nonlinear shape. The negative effect of FDI on GTI is strengthened when the environmental regulation exceeds its threshold. Increasing FDI inflow can inhibit the effect of environmental regulation. Meanwhile, a strict environmental regulation can enhance the inhibiting effect of FDI on GTI. The FDI inflow into high-tech manufacturing sectors has a less negative impact on GTI than the FDI inflow into low-tech sectors in the case of the enhancement of environmental regulation. This study provides some implications for the formulation of environmental regulation and the FDI inflow into China to improve the GTI.

Arsenic trioxide-induced cell apoptosis and cell cycle arrest are potentiated by 1,25-dihydroxyvitamin D3 in human leukemia K562 cells.

The interaction between 1,25-dihydroxyvitamin [1,25(OH)2D3] and vitamin D receptor (VDR) plays a critical role in regulating cell proliferation and programmed cell death. The present study aimed to investigate the effects of 1,25(OH)2D3 in combination with arsenic trioxide (As2O3) on the proliferation and cell cycle of a K562 leukemia cell line. K562 cells were treated with 100 nM 1,25(OH)2D3, 2.5 µM As2O3, and 100 nM 1,25(OH)2D3 combined with 2.5 µM As2O3. Cell proliferation was evaluated by the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt/phenazine ethosulfate method. Cell cycle progression and apoptosis were detected by flow cytometry. The expression levels of genes associated with the cell cycle and apoptosis were analyzed by reverse transcription-quantitative PCR and western blotting analyses. The present findings indicated that combined treatment of 1,25(OH)2D3 and As2O3 led to a significant increase in cytotoxicity, apoptotic cell death and G1 cell cycle arrest when compared to those treated with 1,25(OH)2D3 or As2O3 alone. The downregulation of the Bax/Bcl-2 ratio and decreased survivin expression may be involved in combined treatment-mediated apoptosis. G0/G1 cell cycle arrest induced by combined treatment was associated with the activation of p21 and p27. In addition, the increased expression of VDR was found to participate in the anticancer effect of combination treatment. The data suggested that the combination of 1, 25-(OH)2D3 and As2O3 had clear synergistic effects on the inhibition of K562 cell proliferation, which could provide a novel therapeutic approach for the treatment of acute myeloid leukemia.

[Effects of Decitabine Combined with Bortezomib on the Proliferation of Mantle Cell Lymphoma Cell Lines and Its Underling Mechanisms].

OBJECTIVE: To investigate the effects of decitabine combined with bortezomib on the proliferation of mantle cell lymphoma cell lines (Jeko-1 and Grante519) in vitro and explore the underlying mechanisms. METHODS: Jeko-1 and Grante519 cells were treated with different concentrations of decitabine and/or bortezomib alone and their combination.The cell proliferation was determined by CCK-8 assay. the cell apoptosis were detected by flow cytometry, the mRNA and protein expression levels of genes related with the cell cycle and apoptosis were analyzed by RT-PCR and Western blot respactively. RESULTS: Low dose DAC could significantly inhibit the proliferation and induce apoptosis of Jeko-1 and Grante519 cells which shows a dose-and time-dependent manner. After DAC treatment, caspase 3, BAX and PCDH8 expression levels increased, while BCL-2 and CCND1 expression levels decreased in Jeko-1 and Grante519 cells, but there was no significant difference in NF-κB expression. High dose BTZ could significantly inhibit the proliferation and induce apoptosis of Jeko-1 and Grante519 cells which shows a dose-and time-dependent manner; single drug BTZ could increase the expression level of Caspase 3 and BAX, and decrease the expression level of NF-κB, BCL-2 and CCDN1 in Jeko-1 and Grante519 cells, but there was significant difference in PCDH8 expression level. Compared with single-drug treatment group, DAC combined with BTZ significantly increased the inhibitory rate and apoptotic rate of Jeko-1 and Grante519 cells; PCDH8, Caspase 3 and BAX expression levels significantly increased, and the expression levels of NF-κB, BCL-2 and CCND1 significantly decreased in Jeko-1 and Grante519 cells. CONCLUSION: The combination of DAC and BTZ has obviously synergistic effects on the growth inhibition of Jeko-1 and Grante519 cells which maybe relates with enhancing inbibitory effect on NF-κB signal pathway, down-regulating BAX expression, up-regulating BAX expression as well as increasing cospase 3 expression. This study provides a novel therapeutic approach for mantle cell lymphoma.

Construction of Cu2O/In2O3 Hybrids with p-n Heterojunctions for Enhanced Photocatalytic Performance.

Fabrication of p-n heterojunction materials is an efficient strategy for improving the photocatalytic activity by suppressing the recombination of photogenerated electrons and holes. In this study, a p-n heterojunction consisting of Cu2O and In2O3 was synthesized using the hydrothermal in situ deposition method. The Cu2O/In2O3 hybrid exhibited enhanced performance in the photocatalytic hydrogen evolution reaction and degradation of organic pollutants compared with pure Cu2O and In2O3. The sensitization of In2O3 by Cu2O considerably enhanced the light response of the Cu2O/In2O3 hybrid, and the close contact between In2O3 and Cu2O led to the generation of the p-n heterojunction. The photoelectrochemical properties test proved that the successful fabrication of the p-n heterojunction in the Cu2O/In2O3 hybrid promoted the transfer of photoinduced electrons and holes and inhibited the recombination of photogenerated carriers efficiently. The present strategy provides an efficient approach to improve the performance of photocatalysts through the fabrication of p-n heterojunctions.

Organic solid fluorophores regulated by subtle structure modification: color-tunable and aggregation-induced emission.

Organic solid fluorophores with a tunable emission color have been widely applied in multiple areas. However, rational design and efficient crafting of these fluorophores from a simple core skeleton is still a formidable challenge because of the undesirable concentration quenching emission effect. Herein, we present the development of two series of organic solid fluorophores based on a backbone of p-bis(2,2-dicyanovinyl)benzene. Notably, the introduction of either non-aromatic or aromatic substituents provides fluorophores with a tunable emission color. Moreover, the fluorophores with aromatic substituents exhibit additional unique optical phenomena, such as aggregation-induced emission, polymorphism-dependent emission, and reversible mechanochromic luminescence.

Effective conversion of maize straw wastes into bio-hydrogen by two-stage process integrating H2 fermentation and MECs.

The enhanced H2 production from maize straw had been achieved through the two-stage process of integrating H2 fermentation and microbial electrolysis cells (MECs) in the present work. Several key parameters affecting hydrolysis of maize straw through subcritical H2O were optimized by orthogonal design for saccharification of maize straw followed by H2 production through H2 fermentation. The maximum reducing sugar (RS) content of maize straw reached 469.7 mg/g-TS under the optimal hydrolysis condition with subcritical H2O combining with dilute HCl of 0.3% at 230 °C. The maximum H2 yield, H2 production rate, and H2 content was 115.1 mL/g-TVS, 2.6 mL/g-TVS/h, and 48.9% by H2 fermentation, respectively. In addition, the effluent from H2 fermentation was used as feedstock of MECs for additional H2 production. The maximum H2 yield of 1060 mL/g-COD appeared at an applied voltage of 0.8 V, and total COD removal reached about 35%. The overall H2 yield from maize straw reached 318.5 mL/g-TVS through two-stage processes. The structural characterization of maize straw was also carefully investigated by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), and X-ray diffraction (XRD) spectra.

Direct degradation of cellulosic biomass to bio-hydrogen from a newly isolated strain Clostridium sartagoforme FZ11.

A mesophilic hydrogen-producing strain, Clostridium sartagoforme FZ11, had been newly isolated from cow dung compost acclimated using microcrystalline cellulose (MCC) for at least 30 rounds in an anaerobic bioreactor, and identified by the 16S rDNA gene sequencing, which could directly utilized various carbon sources, especially cellulosic biomass, to produce hydrogen. The maximum hydrogen yields from MCC (10 g/l) and carboxymethyl cellulose (CMC, 10 g/l) were 77.2 and 64.6 ml/g, separately. Furthermore, some key parameters of affecting hydrogen production from raw corn stalk were also optimized. The maximal hydrogen yield and substrate degradation rate from raw corn stalk were 87.2 ml/g and 41.2% under the optimized conditions with substrate concentration of 15 g/l, phosphate buffer of 0.15 M, urea of 6 g/l and initial pH of 6.47 at 35 °C. The result showed that the strain FZ11 would be an ideal candidate to directly convert cellulosic biomass into bio-hydrogen without substrate pretreatment.

Quantitative detection of the human cervical cancer oncogene for monitoring the minimal residual disease in acute leukemia.

The human cervical cancer oncogene (HCCR) has been shown to be over-expressed in some solid tumors, and its function is involved in negative regulation of p53 tumor suppressor gene. However, the roles of HCCR in leukemia remain unclear. The present study is to investigate whether the expression levels of HCCR mRNA are associated with clinical prognosis in patients with acute leukemia (AL) and to explore the potential use as a biomarker for monitoring minimal residual disease (MRD) in AL. The mRNA levels of HCCR1 and HCCR2 were quantified by real-time reverse transcription polymerase chain reaction in bone marrow samples from 80 adult de novo AL patients and 20 normal healthy donors. The expressions of HCCR1 and HCCR2 were significantly higher in patients with acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) than those in healthy donors (P < 0.01), but there was no significant difference between AML and ALL (P > 0.05). Besides white blood cell count, we did not find any significant correlation between HCCR expression and clinical characteristics, such as age, sex, CD34 antigen expression, and response to chemotherapy. HCCR was monitored in 12 cases during remission and/or relapse. Significant reductions of both HCCR1 and HCCR2 mRNA levels were observed in patients who had achieved complete remission after chemotherapy but not in patients with non-responsive. However, an increased HCCR expression was detected in these patients who relapsed. Our findings suggest that HCCR gene is over-expressed in AL patients and may be as a useful biomarker for monitoring MRD in AL.

[Expression of c-fes gene in leukemia cells and its clinical significance].

This study was purposed to investigate the expression of c-fes gene in leukemia patients and its clinical significance. The expression of c-fes mRNA in bone marrow cells from 121 cases of acute and chronic leukemia patients, and the expression of c-fes mRNA in peripheral blood mononuclear cells of 20 normal persons were detected by real time-quantitative reverse transcription polymerase chain reaction (RQ-PCR). The results showed that the level of c-fes mRNA in AML patients was higher than that in normal controls [(48.017 +/- 57.170) x 10(-3) vs (0.152 +/- 0.398) x 10(-3)] (p < 0.0001); but there was no significant differences of level of c-fes mRNA between samples of ALL and normal controls(0.047 +/- 0.068) x 10(-3) vs(0.152 +/- 0.398) x 10(-3) (p>0.05); the level of c-fes mRNA in CML patients was higher than that in normal persons (21.605 +/- 24.818) x 10(-3) vs (0.152 +/- 0.398) x 10(-3) (p < 0.0001). The positive expression rate of c-fes gene in CML-CP patients (80%) was higher than that in CML-AP patients (66.7%) and CML-BP (28.6%) patients. In AML patients, c-fes gene was expressed higher in M(2) (80.77%) and M(3) (92.86%) patients. The remission rate of AML (except M(3))patients who had expression of c-fes gene was 81.08%, which was higher than that of patients with no expression of c-fes gene (40.00%). It is concluded that c-fes gene expression was found in myeloid leukemias, whereas low or no expression in lymphocytic leukemias. The differentiation of myelocytic cells may be related to c-fes gene. All AML (except M(3))patients with high level of c-fes mRNA may get good prognosis.

[Effects of climate change on photosynthesis of Diospyros kaki under different soil moisture conditions].

Taking two-year-old Diospyros kaki as test material, this paper studied the effects of high CO2 concentration (700 micromol x mol(-1)), high temperature (5 degrees C higher than the mean daily temperature); and their combination on the net photosynthesis rate (Pn), evapotranspiration (Tr), stomatal conductance (Gs), water use efficiency (WUE), chlorophyll content, Fv/Fm, and Fv/Fo under different soil moisture conditions, with the ambient air temperature and CO2 concentration (380 micromol x mol(-1)) as the control. Under all test soil moisture conditions, the combination of high CO2 concentration and high temperature decreased the Tr and Gs, but increased the WUE. This combination increased the Pn when the soil moisture content was 75%-85% and 55%-65% of field capacity, but decreased the Pn when the soil moisture content was 35%-45%. High CO2 concentration increased the Pn and WUE but decreased the Gs and Tr under all test soil moisture conditions. The effects of high temperature and its combination with high CO2 concentration on the WUE depended on soil moisture condition, with the WUE increased with increasing soil moisture content. Comparing with the control, high CO2 concentration also increased the leaf Chla, Chlb, Chl (a + b), and Car concentrations and the Fv/Fm and Fv/Fo values, relieved the water stress, and increased the stress-resistance of D. kaki.

[The expression and clinical significance of survivin gene in leukemia].

OBJECTIVE: To investigate the expression of survivin in leukemia and the prognostic significance in acute leukemia (AL). METHODS: The expression of survivin mRNA was measured in 105 AL and 21 chronic myelogenous leukemia (CML) patients with semi-quantity reverse transcription (RT)-PCR. 15 adults were tested as normal control (NC) and K562, NB4, Kg-1alpha, HL-60 cell lines were also tested as positive control. The cell cycle distribution in AL was measured with flow cytometry (FCM) to analyze the relationship between the level of survivin and cell proliferation. RESULTS: The expression of survivin in de novo AL (0.525 +/- 0.460) was higher than that in NC (0.101 +/- 0.187), while it decreased in complete remission (CR) patients (0.280 +/- 0.095). In replased patients (0.935 +/- 0.343), the expression of survivin increased again. There was no difference of the expression between chronic phase of CML (0.279 +/- 0.112) and NC, but in acute phase of CML (0.653 +/- 0.236), the expression was higher than that in NC. The cases with higher level of survivin had significantly higher proliferation indices (mean PI 9.682) than those (mean PI 6.899). In AL patients, the CR rate of survivin positive cases was lower than that of survivin negative cases. CONCLUSIONS: The PI in survivin positive patients was significantly higher than that in survivin negative indicating its impact on proliferation and cleavage. Abnormal expression of survivin genes was related to pathogenesis and progression of AL and it can serve as a marker of relapse and poor prognosis in AL. Overexpression of survivin is a risk factor for CML.

[Expression and significance of X-linked inhibitor of apoptosis protein and its antagonized proteins in acute leukemia].

To investigate the expression and significance of X-linked inhibitor of apoptosis protein (XIAP) and XIAP-associated factor 1 (XAF1) in acute leukemia, the expression of XIAP, XAF1, Smac, and HtrA2 mRNA in the bone marrow aspirates from 87 newly diagnosed AL patients, 23 patients in remission, 6 patients in relapse, and 17 normal controls were detected by means of reverse transcriptase polymerase chain reaction (RT-PCR), and their relationship with clinical therapeutic efficiency was analyzed. The results showed that the expression level of XIAP mRNA in newly diagnosed AL patients (0.990 +/- 0.337) was significantly higher than that in normal controls (0.395 +/- 0.148) (P < 0.01); the positive rate and expression level of XAF1 mRNA in newly diagnosed AL patients (56.32%, 0.246 +/- 0.267) were significantly lower than that in normal controls (100%, 0.964 +/- 0.387) (P < 0.01). In 69 out of 87 newly diagnosed AL patients, efficacy remained evaluable. AL patients with high level of XIAP achieved a lower complete remission (CR) rate than patients with low level of XIAP (54.55% and 86.11%, respectively) (P < 0.01). XAF1 positive patients achieved a higher CR rate than XAF1 negative patients (86.84% and 51.61%, respectively) (P < 0.01). It is concluded that the overexpression of XIAP and negativity of XAF1 may be two adverse prognostic factors in AL patients.

[The study of ribosomal protein l15 cDNA sequences as a molecular marker in the teleostei phylogenetic analysis].

In this study, the full-length cDNAs of RPL15 (ribosomal protein L15) gene were cloned and sequenced from fifteen fishes of five orders under Teleostei. The complete ORF sequences were analyzed for phylogenetic reconstruction for the first time to evaluate the potential of RPL15 gene as a novel marker in resolving teleostean phylogenetic relationships. The resultant NJ, MP and ML trees with Anguilla japonica as the outgroup were generally resolved and largely congruent with the morphology-based tree though some incongruities were observed. The incongruities were then analyzed for some explanations. The results suggested that: (1) RPL15 gene was highly conserved during eukaryotic evolution; (2) RPL15 ORF might be a good phylogenetic marker for resolving teleostean relationships. It might be especially appropriate for the higher-level relationships (such as interordinal), and it was possibly suitable for lower-level relationships as well. The same can be true for other eukaryotes.

Molecular cloning and sequence analysis of full-length growth hormone cDNAs from six important economic fishes.

In this study,the full-length cDNAs of GH (Growth Hormone) gene was isolated from six important economic fishes, Siniperca kneri, Epinephelus coioides, Monopterus albus, Silurus asotus, Misgurnus anguillicaudatus and Carassius auratus gibelio Bloch. It is the first time to clone these GH sequences except E. coioides GH. The lengths of the above cDNAs are as follows: 953 bp, 1 023 bp, 825 bp, 1 082 bp, 1 154 bp and 1 180 bp. Each sequence includes an ORF of about 600 bp which encodes a protein of about 200 amino acid: S. kneri, E. coioides and M. albus GHs of 204 amino acid, S. asotus GH of 200 amino acid, M. anguillicaudatus and C. auratus gibelio GHs of 210 amino acid. Then detailed sequence analysis of the six GHs with many other fish sequences was performed. The six sequences all showed high homology to other sequences, especially to sequences within the same order, and many conserved residues were identified, most localized in five domains. The phylogenetic trees (MP and NJ) of many fish GH ORF sequences (including the new six) with Amia calva as outgroup were generally resolved and largely congruent with the morphology-based tree though some incongruities were observed, suggesting GH ORF should be paid more attention to in teleostean phylogeny.

[Polymorphism in intron 2 of crucian carp GH I gene].

High length and nucleotide polymorphisms in intron2 of GH I gene were detected in 162 individuals,which were from seven wild crucian carp colonies, two goldfish colonies and one Fangzheng crucian carp colony. Using denaturating polyareylamide gel electrophoresis (DPAGE) and single-strand conformation polymorphism (SSCP), seven length variants and 15 haplotypes were identified in these fishes. The length types and haplotypes diversity was 4.32% and 9.26%, respectively. Sequence analysis of the 15 haplotypes indicated the following results: (1) The size of intron2 varied from 243 to 263 bp. In the 15 haplotypes,the average percentages of the four bases (A,T,G and C) were 34.13%, 37.36%, 15.13% and 13.38%, respectively; the frequency of G + C(28.51%) was much lower than that of A + T (71.49%). The GT/AG domain was found in exon-intron junctions,which was the 5' and 3' splice donor and acceptor sites in higher eukaryotic gene introns. The similarity sequence of GTAAGTA was located on the junction between exon2 and intron2. And there existed a richer pyrimidine region (TTTGCCTTTTGTTATC) near the 3' end of intron2. (2) The seven length variants (A, B, C, D, E, F and G) were determined to be 189, 196, 204, 205, 206, 207 and 209 bp, respectively. The polymorphism resulted from the variable repeat number of T (N = 0, 8, 9, 10, 11 and 13) and the difference in one or two motifs deletions of TGAAAAC, TT and GAGTG. (3) Compared the sequences of the 15 haplotypes, 17 substitution sites were observed, of which two were of transversion sites and 15 were of transition sites. Obviously, the transition mutations (88.24%) were more frequent than transversion mutations (11.76%). Analysis of the distributions of the length types, haplotypes and composite genotypes suggested that genetic diversity was varied in different colonies. In the goldfish colonies, only one length type (A), two haplotypes (A1 and A2) and one composite genotype (A1A2) were observed. Two length types (C and D), four haplotypes (C1, C2, D2 and D5) and one composite genotype (C1C2D2D5) presented in the Fangzheng crucian carp colonies. The highest level of genetic diversity was exhibited in the seven wild crucian carp colonies: seven length types (A, B, C, D, E, F and G), 14 haplotypes (A1, A2, A3, B, C1, C2, D1, D2, D3, D4, E1, E2, F and G) and 14 composite genotypes (A1A2A3, A1A2A3B, A1C1C2D1D2D3, A1C1C2D2, A1C1C2D2D3, A1C1C2D3E2, BC1C2D2, BC1C2D3D4, C1C2D2, C1C2D2D3, C1C2D3D4, C1C2D3D4F, C1C2D4, D2E1G) were shared by the seven wild colonies. The numbers of observed length types, haplotypes and genotypes within the wild colonies ranged from 3 to 6, 6 to 10 and 2 to 6, respectively. Whether the length and nucleotide polymorphisms in the intron2 of crucian carp GH I gene were associated with gene expression and gene regulation remained unsolved and required further investigations.

[Relationship between resistance to chemotherapy and expression of breast cancer resistance protein (BCRP) gene in patients with acute leukemia].

In order to investigate the relationship between the expression of breast cancer resistance protein (BCRP) gene and drug resistance in patients with acute leukemia (AL), semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) was used to examine the expression of BCRP mRNA in AL patients and 15 normal subjects. Beta(2)-microglobin (beta(2)-MG) was used as positive reference. BCRP/beta(2)-MG ratio >or= 0.5 was defined as BCRP mRNA positive. The results showed that the positive percentage of BCRP gene expression in newly diagnosed group was 37.6%. The first complete remission rate was 79.3% and 31.6% in BCRP mRNA negative and BCRP mRNA positive patients respectively. The difference was significant (P = 0.001). The expression levels of BCRP mRNA in drug resistance group and drug sensitive group were 0.962 +/- 0.426 and 0.315 +/- 0.296 respectively (P = 0.0001). The expression level of BCRP mRNA in relapsed/refractory group was significantly higher than that in newly diagnosed group (P = 0.0025). The expression level of BCRP gene in normal individuals and long-term survival group was very low and correlated with FAB subtypes. It is concluded that high expression of BCRP mRNA leads to clinical drug resistance and is an unfavorable factor for prognosis of AL patients except acute promyelocytic leukemia.