Exogenous α-calcitonin gene-related peptide attenuates lipopolysaccharide-induced acute lung injury in rats.

α-Calcitonin gene-related peptide (α-CGRP) is a 37 amino-acid neuropeptide that is primarily released from C-type sensory neurons. α-CGRP exerts multiple modulatory effects on immune responses and visceral organ function, but the role of exogenous α-CGRP in lipopolysaccharide (LPS)-induced acute lung injury (ALI) has remained to be elucidated. Forty-eight rats were randomized to receive continuous intraperitoneal infusion of α-CGRP (0.4 µg/kg/min) or normal saline for 30 min, followed by intratracheal injection of 0.5 mg/kg LPS or saline. There were four groups of animals: The saline-saline (S-S) group; the saline-α-CGRP (S-C) group; the LPS-saline (L-S) group and the LPS-α-CGRP (L-C) group. Mean arterial pressure and arterial blood gases were assessed prior to α-CGRP and LPS administration and every hour following LPS treatment. After 4 h, bronchoalveolar lavage was performed and used to assess total cell count and levels of tumor necrosis factor-α, interleukin-1ß, intracellular cell adhesion molecule 1 and macrophage inflammatory protein 2. Lung tissue was also collected for assessing wet-to-dry (W/D) ratio, histology and Evans blue (EB) dye extravasation. Pulmonary α-CGRP concentration and α-CGRP receptor expression were also examined, and inducible cyclic adenosine monophosphate early repressor (ICER) and TNF-α mRNA expression levels were measured. Treatment with exogenous α-CGRP improved oxygenation during LPS-induced ALI. Correspondingly, histological injury, total cell count, inflammatory cytokine levels, W/D ratio and EB dye extravasation were also significantly reduced. α-CGRP receptor 1 expression was noted in pulmonary endothelial cells and alveolar macrophages and α-CGRP receptor expression levels were decreased during ALI, whereas pulmonary α-CGRP expression was continuously increased. Furthermore, exogenous α-CGRP induced upregulation of ICER during LPS-induced ALI. In conclusion, exogenous α-CGRP improved oxygenation and ameliorated lung damage in LPS-induced ALI, and these effects were associated with the upregulation of ICER.

Surveillance of childhood blood lead levels in Chengdu, China in 2010-2011.

INTRODUCTION: Lead poisoning has been receiving great attention around the world. The Child Hygiene Cooperation Center of the World Health Organization in China has been conducting investigations to monitor blood lead levels (BLLs) from as early as 2004. However, only several lead poisoning studies have been conducted in China since August 2009. The aim of the present study was to investigate the BLLs in children aged < 7 years and to analyse the risk factors of high BLLs in Chengdu, China. METHODS: Questionnaires were distributed to children in Chengdu from 2010 to 2011. A total of 2,271 children were included in this study - 1,157 received BLL tests in 2010 and the remaining received the tests in 2011. BLL was measured using a tungsten atomiser absorption spectrophotometer. RESULTS: The mean BLL of the 2,271 children was 6.2 µg/dL and 2.03% of the children had BLLs ≥ 10 µg/dL. Mean BLL seemed to increase with age. Unhygienic habits (e.g. not washing hands frequently, biting of toys and pencils), history of pica, use of coal and residence in an industrial zone were found to be the main risk factors contributing to high BLL (p < 0.05). Children with high BLLs have a higher risk of manifesting anorexia and/or abdominal pain as compared to those with low BLLs (p < 0.05). CONCLUSION: The mean BLL of children in Chengdu (i.e. 6.2 µg/dL) was found to be higher than that of children in developed countries. Childhood lead poisoning remains a public health problem.

Elevated serum ferritin levels in patients with hematologic malignancies.

PURPOSE: To retrospectively analyze variability and clinical significance of serum ferritin levels in Chinese patients with hematologic malignancies. MATERIALS AND METHODS: Serum ferritin were measured by radioimmunoassay, using a kit produced by the Beijing Institute of Atomic Energy. Patients with hematologic malignancies, and treated in the Department of Hematology in Nanjing First Hospital and fulfilled study criteria were recruited. RESULTS: Of 473 patients with hematologic malignancies, 262 patients were diagnosed with acute leukemia, 131 with lymphoma and 80 with multiple myeloma. Serum ferritin levels of newly diagnosed and recurrent patients were significantly higher than those entering complete remission stage or in the control group (p<0.001). CONCLUSIONS: Serum ferritin lever in patients with hematologic malignancies at early stage and recurrent stage are significantly increased, so that detection and surveillance of changes of serum ferritin could be helpful in assessing conditions and prognosis of this patient cohort.

[Methylation of FHIT gene promoter region in DNA from plasma of patients with myelodysplastic syndromes and demethylating effect of decitabine].

This study was aimed to detect the methylation status of FHIT gene promoter region in the DNA from plasma of patients with myelodysplastic syndrome (MDS), and to investigate the demethylating effect of decitabine. Methylation-specific PCR method was used to detect the methylation status of FHIT gene promoter region in the DNA from plasma of 4 patients with MDS before and after treatment with decitabine plus semis CAG therapy (among them, 1 case of newly diagnosed MDS, 3 cases progressed into acute leukemia). The results indicated that 3 cases were found to have an increased methylation in the promoter region. After treatment with decitabine plus semis CAG, increased methylation was reversed in 2 cases. In 4 cases, 2 cases displayed clinical response. It is concluded that FHIT gene hypermethylation is associated with MDS pathogenesis. Decitabine has demethylating effect on the FHIT gene hypermethylation of plasma from MDS patients. Detecting the methylation status of FHIT gene in DNA from plasma may play a role in MDS auxiliary diagnosis or prognosis.

Phase II study on voriconazole for treatment of Chinese patients with malignant hematological disorders and invasive aspergillosis.

OBJECTIVE: To investigate the efficacy and safety of voriconazole in treating Chinese patients with hematological malignancies and invasive aspergillosis. METHODS: From March 2007 to April 2012, patients with diagnoses confirmed by CT, GM test and/or PCR assays, were recruited into this study. Aspergillosis of all patients were treated with voriconazole 6 mg/kg intravenous infusion (iv) every 12 h for 1 day, followed by 4 mg/kg IV every 12 h for 10-15 days; Then, switch to oral administration that was 200 mg every 12 h for 4-12 weeks. Efficacy and safety were evaluated according to Practice Guideline of Infectious Diseases Society of America. RESULTS: The overall response rate of 38 patients after voriconazole treatment was 81.6%. The median time to pyretolysis was 4.5 days. Treatment related side effects were mild and found in only 15.8% of cases. No treatment related deaths occurred. CONCLUSIONS: Voriconazole can considered to be a safe and effective front-line therapy to treat patients with hematological malignancies and invasive aspergillosis. Alternatively it could be used as a remedial treatment when other antifungal therapies are ineffective.

[Detection of molecular cytogenetic abnormalities in 30 patients with chronic lymphocytic leukemia by fluorescence in situ hybridization].

This study was aimed to investigate the significance of interphase fluorescence in situ hybridization (FISH) in detecting +12, del (13q14), p53 and atm gene deletion in chronic lymphocytic leukemia (CLL). FISH and a panel of probes (CEP 12, LSI D13S319, LSI p53, LSI atm) were used to detect molecular cytogenetic abnormalities in 30 patients with CLL. Cytogenetic aberrations and their relation with some other prognostic factors (peripheral lymphocyte count, Binet stage, LDH level, ZAP-70 and so on) were analyzed. The results indicated that out of the 30 CLL patients, molecular cytogenetic aberrations were found in 19 (63.3%) cases and 7 (23.3%) patients showed more than two kinds of abnormalities. The most frequent abnormality detected was del (13q14) (43.3%), followed by trisomy of chromosome 12 (23.3%), del (atm) (13.3%) and del (p53) (10.0%). There were no significant differences between molecular cytogenetic aberrations and sex, age, Binet stage, peripheral lymphocyte count, or the serum levels of lactate dehydrogenase (LDH), beta(2)-microglobulin (beta(2)-MG), or ZAP-70. The incidence of atm gene deletion was higher in the group of CD38 high expression than that in the group of low expression (p = 0.035). It is concluded that FISH is a rapid and sensitive technique in analysing molecular cytogenetic abnormalities, but its prognostic significance in CLL needs to further investigate.

Protease-catalyzed synthesis of a precursor dipeptide, Z-Asp-Val-NH2 of thymopentin, in organic solvents.

The protease-catalyzed, kinetically controlled synthesis of a precursor dipeptide, Z-Asp-Val-NH(2) of thymopentin (TP-5), in organic solvents was studied. Z-Asp-OMe and Val-NH(2) were used as the acyl donor and the nucleophile, respectively. An industrial alkaline protease alcalase was used to catalyze the synthesis of the target dipeptide in water-organic cosolvent systems. The conditions of the synthesis reaction were optimized by examining the effects of several factors, including organic solvents, water content, temperature, pH, and reaction time on the yield of Z-Asp-Val-NH(2). The optimum conditions using alcalase as the catalyst are pH 10.0, 35 degrees C, in acetonitrile/Na(2)CO(3)-NaHCO(3) buffer system (9:1, V/V), reaction time 5 h, with a yield of 63%. The dipeptide product was confirmed by LC- MS.

Synthesis of a precursor dipeptide of thymopentin in organic solvents by an enzymatic method.

The protease-catalyzed, kinetically controlled synthesis of a precursor dipeptide of thymopentin(TP-5), Z-Arg-Lys-NH2 in organic solvents was studied. Z-Arg-OMe was used as the acyl donor and Lys-NH2 was used as the nucleophile. An industrial alkaline protease alcalase and trypsin were used to catalyze the synthesis of the target dipeptide in water-organic cosolvent systems. The conditions of the synthesis reaction were optimized by examining the effects of several factors, including organic solvents, water content, temperature, pH, and reaction time on the yield of Z-Arg-Lys-NH2. The optimum conditions using alcalase as the catalyst are pH 10.0, 35 degrees C, in acetonitrile/DMF/Na2CO3-NaHCO3 buffer system (80:10:10, V/V), 6 h, with the dipeptide yield of 71.1%. Compared with alcalase, the optimum conditions for trypsin are pH 8.0, 35 degrees C, in ethanol/Tris-HCl buffer system (80:20, V/V), 4 h, with the dipeptide yield of 76.1%.

Alcalase-catalysed synthesis of the precursor tetrapeptide N-benzoylarginylglycylaspartylserinamide (Bz-RGDS-NH2) of the cell-adhesion peptide arginylglycylaspartylserine (RGDS).

In the present study, a precursor tetrapeptide Bz-RGDS-NH2 (N-benzoylarginylglycylaspartylserinamide) of cell-adhesion peptide RGDS (arginylglycylaspartylserine) was synthesized by a novel route. First of all, the precursor tripeptide GDS-NH2 (glycylaspartylserinamide) was synthesized by a chemical method only using aspartic acid and serine at gram scale in four steps. The linkage of the fourth amino acid Bz-Arg-OEt (N-benzoyl-L-arginine ethyl ester) to GDS-NH2 was completed by an enzymatic method under kinetic control in water-miscible organic media. An industrial alkaline protease, Alcalase, with a wide substrate specificity, was used as the catalyst. The effects of organic solvents, pH value, reaction temperature, water content and molar ratio of substrates on the yield of Bz-RGDS-NH2 synthesis were examined. The optimum reaction conditions were found to be pH 10.0, 35 degrees C, 8 h, in an acetonitrile/(Na2CO3/NaHCO3) buffer system (93:7, v/v) with a maximal yield of 65.2%. We found that secondary hydrolysis of the peptide product did not take place in these water-miscible organic solvents.

Preparation of corn (Zea mays) peptides and their protective effect against alcohol-induced acute hepatic injury in NH mice.

CPS [corn (Zea mays) peptides] were prepared from corn gluten meal by proteolysis with alcalase, an alkaline protease. The molecular-mass distribution of CPS is from 200 to 1000 Da as determined by MS. The amino acid composition of CPS was also analysed by HPLC. CPS contains almost no free amino acids. The protective effect of CPS against acute hepatic injuries induced by alcohol was verified in NH mice that were fed with different dosages of CPS for 30 days and subsequently given an acute dose of alcohol orally. As a result, CPS reduced both hepatic malondialdehyde and triacylglycerol levels, along with enhanced hepatic GSH (glutathione) levels, relative to the control. Hepatic histological changes were also observed. The result indicates that CPS is capable of attenuating ethanol-induced hepatic injury. The effect of CPS on removing superoxide anion in vitro was also studied as an additional proof that CPS is capable of abating hepatic superoxidant stress.

Clinical observation on treatment of chronic aplastic anemia by Shengxuening and cyclosporin A.

OBJECTIVE: To explore the therapy to further elevate the efficacy of the treatment of chronic aplastic anemia (CAA). METHODS: Forty-five patients with CCA were assigned into two groups, the 26 patients in the treated group were treated by Shengxuening (a Chinese herbal preparation) and cyclosporin A (CsA), and the 19 patients in the control group were treated with androgen alone, with the therapeutic course lasting for over 3 months. Changes of peripheral blood picture, and the colony productivity of burst forming unit-erythroid (BFU-E), colony forming unit-erythroid (CFU-E) and colony forming unit-granulocyte macrophage (CFU-GM) in bone marrow were observed before and after 3 months treatment. The amount of erythrocyte and platelet infusion, frequency of infection, condition of hemorrhage and relevant death were also observed. The follow-up study was conducted for over half a year. RESULTS: The total effective rate in the treated group was 84.6%, which was significantly higher than that in the control group (52.6%, P < 0.05). Levels of hemoglobin, reticulocyte, neutrophil and platelet increased after treatment in the treated group, as compared with those before treatment, with significant difference (P < 0.05), and the colony productivity of BFU-E, CFU-E and CFU-GM in bone marrow also got significantly increased (P < 0.01), and showed significant difference from those in the control group (P < 0.05). CONCLUSION: Shengxuening-assisting CsA therapy is an effective measure for treatment of CAA.

New synthetic route for RGD tripeptide.

A new route was employed to synthesize RGD. First, Gly-Asp dipeptide was synthesized by a novel chemical method in two steps, including chloroacetylation of L-aspartic acid and ammonolysis of chloroacetyl L-aspartic acid. Second, Nalpha-Z- L-Arginine was reacted with Gly-Asp to synthesize RGD by the N-carboxyanhydride method. Less protected amino acids were used in this synthesis. This method possessed advantages of low cost, simplicity, and rapidity with a reasonable yield of 62% calculated from arginine. In addition, compared with the above method, a conventional solid phase method was also used to synthesize RGD, the yield was 75% calculated from the first amino acid anchored to resin.

Synthesis of tetrapeptide Bz-RGDS-NH2 by a combination of chemical and enzymatic methods.

The tetrapeptide Bz-Arg-Gly-Asp-Ser-NH(2) (Bz-RGDS-NH(2)) was successfully synthesized by a combination of chemical and enzymatic methods in this study. Firstly, the precursor tripeptide Gly-Asp-Ser-NH(2) (GDS-NH(2)) was synthesized by a novel chemical method in four steps including chloroacetylation of l-aspartic acid, synthesis of chloroacetyl l-aspartic acid anhydride, the synthesis of ClCH(2)COAsp-SerOMe and ammonolysis of ClCH(2)COAsp-SerOMe. Secondly, lipase (PPL) was used to catalyze the formation of Bz-RGDS-NH(2) in aqueous water-miscible organic cosolvent systems using Bz-Arg-OEt as the acyl donor and GDS-NH(2) as the nucleophile. The optimum conditions were Bz-Arg-OEt 50 mM; GDS-NH(2) 400 mM; 10 degrees C, 0.1M phosphate buffer, pH 7.5; 60% DMF or 58% DMSO, PPL: 10 mg ml(-1) with the maximum yields of the tetrapeptide of 73.6% for DMF and 70.4% for DMSO, respectively. The secondary hydrolysis of the tetrapeptide product did not take place due to the absence of amidase activity of lipase.

Synthesis of a precursor dipeptide of RGDS (Arg-Gly-Asp-Ser) catalysed by the industrial protease alcalase.

Synthesis of Bz-Arg-Gly-NH(2) (N-benzoylargininylglycinamide) [a precursor dipeptide of RGDS (Arg-Gly-Asp-Ser)] catalysed by protease in water/organic co-solvent systems was studied. Starting substrates were N-benzoyl-L-arginine ethyl ester hydrochloride (acyl donor) and glycinamide (nucleophile). Acetonitrile was selected as the organic solvent. Alcalase, an industrial alkaline protease, was applied to the synthesis of the target dipeptide. The conditions of the synthesis reaction were optimized by examining the effects of several factors, including water content, temperature, pH, molar ratio of the substrates and reaction time, on the yield of Bz-Arg-Gly-NH(2). The optimum conditions were established to be pH 10.0, 45 degrees C, in acetonitrile/0.1 M Na(2)CO(3)/NaHCO(3) buffer system (90:10, v/v) for 1 h with a dipeptide yield of 82.9%.

Alcalase-catalyzed, kinetically controlled synthesis of a precursor dipeptide of RGDS in organic solvents.

The protease-catalyzed, kinetically controlled synthesis of a precursor dipeptide of RGDS, Z-Asp-Ser-NH2 in organic solvents was studied. Alcalase, an industrial alkaline protease, was used to catalyze the synthesis of the target dipeptide in water-organic cosolvents systems with Z-Asp-OMe as the acyl donor and Ser-NH2 as the nucleophile. Acetonitrile was selected as the organic solvent from acetonitrile, ethanol, methanol, DMF, DMSO, ethyl acetate, 2-methyl-2-propanol, and chloroform tested under the experimental conditions. The conditions of the synthesis reaction were optimized by examining the effects of several factors, including water content, temperature, pH, and reaction time on the Z-Asp-Ser-NH2 yields. The optimum conditions are pH 10.0, 35 degrees C, in acetonitrile/Na2CO3-NaHCO3 buffer system (85:15, v/v), 6 h, with a dipeptide yield of 75.5%.

Chemo-enzymatic synthesis of tripeptide RGD diamide in organic solvents.

The tripeptide BzArgGlyAsp(NH(2))(2) was synthesized by a combination of chemical and enzymatic methods in this study. First of all, GlyAsp(NH(2))(2) was synthesized by a novel chemical method in three steps including chloroacetylation of L-aspartic acid, esterification of chloroacetyl L-aspartic acid and ammonolysis of chloroacetyl L-aspartic acid diethyl ester. Secondly, kinetically controlled synthesis of BzArgGlyAsp(NH(2))(2) catalyzed by trypsin in organic solvent was conducted. The optimum conditions are pH 8.0, 30 degrees C in ethanol/Tris-HCl buffer system (85:15, v/v) for 80 min in the maximum yield of 74.4%.

Trypsin-catalyzed kinetically controlled synthesis of a precursor dipeptide of thymopentin in organic solvents, using a free amino acid as nucleophile.

Trypsin-catalyzed, kinetically controlled synthesis of a precursor, dipeptide of thymopentin (TP-5), Bz-Arg-Lys-OH (or-OEt) in organic solvents was studied. Bz-Arg-OEt was used as the acyl donor and Lys-OH and Lys-OEt were used as the nucleophiles. Ethanol was selected as the organic solvent from ethanol, methanol, acetonitrile, and ethyl acetate tested under the experimental conditions. As expected, Lys-OEt is not a suitable nucleophile in trypsin-catalyzed reaction, due to its competition with the protective Arg-OEt as acyl donor for the active site of trypsin, while Lys-OH does not have this problem. The optimal reaction condition for the synthesis of Bz-Arg-Lys-OH was set up as 20% Tris-HCl buffer, pH 8.0, 35 degrees C for 6 h with the yield of 52.5%, or for 18-24 h with the yield of about 60%.

[Detection of residual leukemic cells after complete remission of patients with acute myeloid leukemia].

The aim was to study minimal residual disease (MRD) in blood and bone marrow after complete remission of patients with acute myeloid leukemia (AML) and explore the role of MRD in detecting relapse of acute myeloid leukemia. The blood and bone marrow samples from 33 AML patients who had been in complete remission were determined for residual leukemic cells (RLC) with flow cytometry. The results showed that RLC in AML group of complete remission was higher than that of normal group both in blood by (4.7518 +/- 4.1537)% vs (0.4835 +/- 0.2005)% and bone marrow by (17.9082 +/- 20.4819)% vs (0.7285 +/- 0.2209)%, while the RLC in relapsed group was higher than that in non-relapsed group both in blood by (2.233 +/- 1.5923)% vs (10.2369 +/- 9.4714)% and bone marrow by (4.779 +/- 3.0336)% vs (38.0685 +/- 19.4295)%. In conclusion, early detection of leukemic residual cells with flow cytometry contributes to treatment of relapse in time.

Lipase-catalyzed synthesis of precursor dipeptides of RGD in aqueous water-miscible organic solvents.

PPL-catalyzed synthesis of the precursor dipeptides of RGD as a cellular adhesion factor, Benzyl-Arg-Gly-NH2 and CBZ-Gly-Asp-NH2, was conducted in water-organic cosolvents systems. Five water-miscible organic solvents, which have some advantage over the water-immiscible organic solvent systems or the anhydrous organic solvent systems used often in protease-catalyzed synthesis of a peptide bond, were tested. The reaction condition of PPL-catalyzed synthesis of the dipeptides was optimized by examining the main factors affecting the product yield. The optimal reaction condition for the synthesis of Benzyl-Arg-Gly-NH2 was set up as pH 8.0, 15 degrees C in 40% MeOH for 10 h with the maximum yield of 73.6%. The optimum condition for the synthesis of CBZ-Gly-Asp-NH2 was pH 7.0, 15 degrees C in 50% MeOH for 10h with the maximum yield of 67.0%.