Objective To evaluate the engraftment state after allogeneic peripheral blood stem cell transplantation(Allo-PBSCT)in 2 exposed patients in Shandong “10?21” radiation accident.Methods The dynamic cytogenetic analysis was performed by chromosome aberration(CA)analysis,R-band karyotyping analysis and micronucleus(MN)assay of peripheral blood and/or bone marrow samples of 2 patients(A and B).Results The results showed that the transplantation was successful(complete chimerism),and the engraftment state was stable.No difference was observed between the results of cytogenetic analysis from peripheral blood samples and bone marrow samples.These results were in fairly good accordance with those results by short tandem repeats-polymerase chain reaction(STR-PCR)and clinical manifestations.Conclusions The results suggested that the dynamic observations of dicentric and ring aberrations and MN can be used as an indication to estimate the engraftment state after Allo-PBSCT,but only for the patient with acute radiation sickness.Moreover,the cytogenetic analysis performed on bone marrow samples is better than that on peripheral blood samples when the lymphocytes were reduced sharply in peripheral blood.

Objective A biological dose estimation by cytokinesis-block micronucleus(CBMN)assay was performed on 2 patients exposed to accidental radiation.Method Micronucleus assay and dose estimation by CBMN in peripheral blood and bone marrow were performed in 2 patients(A and B)suffering from radiation sickness.Results No binucleated lymphocytes were observed in peripheral blood of 2 patients.The results of bone marrow examination showed that only one binucleated cell was discovered in patient A.The rough estimation of radiation dose of A was above 20Gy according to the degree of reduction of binucleated cell in our previous study.The micronucleus frequency was 2.42 per cell and the dose estimation was 8.7(8.0-9.4)Gy for patient B.These results were approximately in accordance with that of dose estimations by physical method,chromosome aberration analysis,ESR method,and clinical manifestations.Conclusion MN assay is simple,quick and accurate.It is a reliable biodosimetry in addition to chromosome aberration analysis.

To further investigate the mechanisms of action of icariin (ICA), we assessed the effects of ICA on the in vitro formation of cGMP and cAMP in isolated rabbit corpus cavernosum. Isolated segments of rabbit corpus cavernosum were exposed to increasing concentrations of ICA and the dose-dependent accumulation of cGMP and cAMP was determined in the tissues samples by means of 125I radioimmunoassay. Responses of the isolated tissues preparations to ICA were compared with those obtained with the reference compounds sildenafil (Sild). Furthermore, the effects of ICA on the mRNA expression of specific cGMP-binding phosphodiesterase type V (PDE5) in rat penis were also observed. After incubation with ICA for 6 h or 14 h respectively, the levels of PDE5 mRNA were examined by reverse transcriptase polymerase chain reaction (RT-PCR). The results showed that ICA increased cGMP concentrations directly (P < 0.05), but there was no significant effect on cAMP concentrations (P > 0.05). In the presence of sodium nitroprusside (SNP), a stimulatory agent of cGMP, both ICA and Sild increased cGMP concentrations with increasing dose (P < 0.01). Their EC50 was 4.62 (ICA) and 0.42 (Sild) micromol/L respectively. Under the same condition, ICA and Sild unaltered cAMP level significantly (P > 0.05). There were PDE5A1 and PDE5A2 mRNA expressions in rat corpus cavernosum with PDE5A2 being the dominant isoform. ICA could obviously inhibit these two isoforms mRNA expression in rat penis, and decrease PDE5A1 more pronouncedly (P < 0.01). The present study indicated that the aphrodisiac mechanisms of icariin involved the NO-cGMP signal transduction pathway, with increasing cGMP levels in the corpus cavernosum smooth muscle. The inhibitory effect of icariin on PDE5 mRNA expression, especially on PDE5A1, might account for its molecular mechanisms for its long-term activity.

In order to further investigate the mechanisms of action of berberine (Ber), we assessed the effects of Ber on the mRNA expression of nitric oxide synthases (NOS) in rat corpus cavernosum. After incubation with Ber for 1 or 3 h respectively, the levels of NOS mRNA were examined by reverse transcription polymerase chain reaction (RT-PCR). Our results showed that there were iNOS and eNOS mRNA expressions in rat corpus cavernosum. Ber enhanced eNOS mRNA expression in rat penis, but exhibited no effect on the expression of iNOS mRNA (P > 0.05). The present study indicated that the relaxation of Ber involved the NO-cGMP signal transduction pathway. The enhancing effect of Ber on eNOS mRNA expression might associated with its relaxation of corpus cavernosum.
Berberine/*pharmacology ; Connective Tissue/physiopathology ; Nitric Oxide Synthase/*biosynthesis ; Nitric Oxide Synthase/genetics ; Nitric Oxide Synthase Type I/biosynthesis ; Nitric Oxide Synthase Type I/genetics ; Nitric Oxide Synthase Type III/biosynthesis ; Nitric Oxide Synthase Type III/genetics ; Penile Erection/*physiology ; Penis/*metabolism ; Penis/physiology ; RNA, Messenger/biosynthesis ; RNA, Messenger/genetics

In order to further investigate the mechanisms of action of berberine (Ber), we assessed the effects of Ber on the mRNA expression of nitric oxide synthases (NOS) in rat corpus cavernosum. After incubation with Ber for 1 or 3 h respectively, the levels of NOS mRNA were examined by reverse transcription polymerase chain reaction (RT-PCR). Our results showed that there were iNOS and eNOS mRNA expressions in rat corpus cavernosum. Ber enhanced eNOS mRNA expression in rat penis, but exhibited no effect on the expression of iNOS mRNA (P＞0.05). The present study indicated that the relaxation of Ber involved the NO-cGMP signal thansduction pathway. The enhancing effect of Ber on eNOS mRNA expression might associated with its relaxation of corpus cavernosum.

To further investigate the mechanisms of action of icariin (ICA), we assessed the effects of ICA on the in vitro formation of cGMP and cAMP in isolated rabbit corpus cavernosum. Isolated segments of rabbit corpus cavernosum were exposed to increasing concentrations of ICA and the dose-dependent accumulation of cGMP and cAMP was determined in the tissues samples by means of 125I radioimmunoassay. Responses of the isolated tissues preparations to ICA were compared with those obtained with the reference compounds sildenafil (Sild). Furthermore, the effects of ICA on the mRNA expression of specific cGMP-binding phosphodiesterase type V (PDE5) in rat penis were also observed. After incubation with ICA for 6 h or 14 h respectively, the levels of PDE5 mRNA were examined by reverse transcriptase polymerase chain reaction (RT-PCR). The results showed that ICA increased cGMP concentrations directly (P＜0.05), but there was no significant effect on cAMP concentrations (P＞0.05). In the presence of sodium nitroprusside (SNP), a stimulatory agent of cGMP,both ICA and Sild increased cGMP concentrations with increasing dose (P＜0.01). Their EC50 was 4.62 (ICA) and 0.42 (Sild) μmol/L respectively. Under the same condition, ICA and Sild unaltered cAMP level significantly (P＞0.05). There were PDE5A1 and PDE5A2 mRNA expressions in rat corpus cavernosum with PDE5A2 being the dominant isoform. ICA could obviously inhibit these two isoforms mRNA expression in rat penis, and decrease PDE5A1 more pronouncedly (P＜ 0.01). The present study indicated that the aphrodisiac mechanisms of icariin involved the NO-cGMP signal transduction pathway, with increasing cGMP levels in the corpus cavernosum smooth muscle. The inhibitory effect of icariin on PDE5 mRNA expression, especially on PDE5A1, might account for its molecular mechanisms for its long-term activity.

Background and objectives:Hepatic steatosis and inflammation are key characteristics of non-alcoholic fatty liver disease(NAFLD).However,whether and how hepatic steatosis and liver inflammation are differentially regulated remains to be elucidated.Considering that disruption of 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3(Pfkfb3/iPfk2)dissociates fat deposition and inflammation,the present study examined a role for Pfkfb3/iPfk2 in hematopoietic cells in regulating hepatic steatosis and inflammation in mice. Methods:Pfkfb3-disrupted(Pfkfb3+-)mice and wild-type(WT)littermates were fed a high-fat diet(HFD)and examined for NAFLD phenotype.Also,bone marrow cells isolated from Pfkfb3+/-mice and WT mice were differentiated into macrophages for analysis of macrophage activation status and for bone marrow transplantation(BMT)to generate chimeric(WT/BMT-Pfkfb3+/-)mice in which Pfkfb3 was disrupted only in hematopoietic cells and control chimeric(WT/BMT-WT)mice.The latter were also fed an HFD and examined for NAFLD phenotype.In vitro,hepatocytes were co-cultured with bone marrow-derived macrophages and examined for hepatocyte fat deposition and proinflammatory responses.Results:After the feeding period,HFD-fed Pfkfb3+/-mice displayed increased severity of liver inflam-mation in the absence of hepatic steatosis compared with HFD-fed WT mice.When inflammatory activation was analyzed,Pfkfb3+/-macrophages revealed increased proinflammatory activation and decreased anti-proinflammatory activation.When NAFLD phenotype was analyzed in the chimeric mice,WT/BMT-Pfkfb3+/-mice displayed increases in the severity of HFD-induced hepatic steatosis and inflammation compared with WT/BMT-WT mice.At the cellular level,hepatocytes co-cultured with Pfkfb3+/-macrophages revealed increased fat deposition and proinflammatory responses compared with hepatocytes co-cultured with WT macrophages. Conclusions:Pfkfb3 disruption only in hematopoietic cells exacerbates HFD-induced hepatic steatosis and inflammation whereas the Pfkfb3/iPfk2 in nonhematopoietic cells appeared to be needed for HFD feeding to induce hepatic steatosis.As such,the Pfkfb3/iPfk2 plays a unique role in regulating NAFLD pathophysiology.