[Interleukin-9 promotes the activation of hepatic stellate cells in mice infected with Schistosoma japonicum].

OBJECTIVE: To investigate the effects of interleukin-9 (IL-9) in the activation of hepatic stellate cells (HSCs) in mice infected with Schistosoma japonicum. METHODS: Primary HSCs were isolated from mice 7 weeks post-infection with S. japonicum using the in situ liver perfusion and density gradient centrifugation, and cultured in vitro. HSCs were randomly assigned to the PBS control group and IL-9 stimulation group (stimulation with 20 ng/mL IL-9). HSCs were harvested 48 h and 72 h poststimulation, and the expression of α-smooth muscle actin (α-SMA), type I collagen (Col I) and type III collagen (Col III) was determined in HSCs using Western blotting. RESULTS: Following stimulation with 20 ng/mL IL-9 for 48 h, the expression of α-SMA [(0.87 ± 0.02) vs. (0.69 ± 0.01); t = 17.39, P < 0.01], Col I [(0.74 ± 0.02) vs. (0.65 ± 0.01); t = 9.56, P < 0.01] and Col III [(0.94 ±0.04) vs. (0.75 ± 0.03); t = 6.15, P < 0.01] was significantly greater in HSCs in the IL-9 stimulation group than in the PBS control group. Following stimulation with 20 ng/mL IL-9 for 72 h, the expression of α-SMA was significantly greater in HSCs in the IL-9 stimulation group than in the PBS control group[(0.76 ± 0.02) vs. (0.58 ± 0.02); t = 12.52, P < 0.01]; however, there were no significant differences between the two groups in terms of Col I [(0.68 ± 0.02) vs. (0.66 ± 0.02); t = 1.15, P > 0.05] or Col III expression [(0.75 ± 0.01) vs. (0.72 ± 0.02); t = 2.22, P > 0.05]. CONCLUSIONS: IL-9 promotes the activation of HSCs in mice infected with S. japonicum.

Prolonged sympathetic innervation of sensory neurons in rat thoracolumbar dorsal root ganglia during chronic colitis.

BACKGROUND: Peripheral irritation-induced sensory plasticity may involve catecholaminergic innervation of sensory neurons in the dorsal root ganglia (DRG). METHODS: Catecholaminergic fiber outgrowth in the thoracolumbar DRG (T13-L2) was examined by tyrosine hydroxylase (TH) immunostaining, or by sucrose-potassium phosphate-glyoxylic acid histofluorescence method. TH level was examined by Western blot. Colonic afferent neurons were labeled by retrograde neuronal tracing. Colitis was induced by intracolonic instillation of tri-nitrobenzene sulfonic acid (TNBS). KEY RESULTS: The catecholaminergic fibers formed 'basket-like' structures around the DRG cells. At 7 days following TNBS treatment, the number of DRG neurons surrounded by TH-immunoreactive fibers and the protein levels of TH were significantly increased in T13, L1, and L2 DRGs (two- to threefold, P < 0.05). The DRG neurons that were surrounded by TH immunoreactivity were 200 kDa neurofilament-positive, but not isolectin IB4-positive or calcitonin gene-related peptide-positive. The TH-immunoreactive fibers did not surround but adjoin the specifically labeled colonic afferent neurons, and was co-localized with glial marker S-100. Comparison of the level of TH and the severity of colonic inflammation showed that following TNBS treatment, the degree of colonic inflammation was most severe at day 3, subsided at day 7, and significantly recovered by day 21. However, the levels of TH in T13-L2 DRGs were increased at both 3 days and 7 days post TNBS treatment and persisted up to 21 days (two- to fivefold increase, P < 0.05) as examined. CONCLUSIONS & INFERENCES: Colonic inflammation induced prolonged catecholaminergic innervation of sensory neurons, which may have relevance to colitis-induced chronic visceral hypersensitivity and/or referred pain.

Sensitive and rapid detection of Schistosoma japonicum DNA by loop-mediated isothermal amplification (LAMP).

In this study, a loop-mediated isothermal amplification (LAMP) assay was established to detect Schistosoma japonicum DNA in faecal and serum samples of rabbits, and serum samples of humans infected with S. japonicum. This LAMP assay was based on the sequence of highly repetitive retrotransposon SjR2, and was able to detect 0.08 fg S. japonicum DNA, which is 10(4) times more sensitive than conventional PCR. The LAMP assay was also highly specific for S. japonicum and able to detect S. japonicum DNA in rabbit sera at 1 week p.i. Following administration of praziquantel, detection of S. japonicum DNA in rabbit sera became negative at 12 weeks post-treatment. These results demonstrated that LAMP was effective for early diagnosis of, and evaluation of therapy effectiveness for, S. japonicum infection. Both PCR and LAMP assays were then used to detect S. japonicum DNA in 30 serum samples from S. japonicum-infected patients and 20 serum samples from healthy persons. The percentage sensitivity of LAMP was 96.7%, whereas that of PCR was only 60%, indicating that LAMP was more sensitive than conventional PCR for clinical diagnosis of schistosomiasis cases in endemic areas. The established LAMP assay should provide a useful and practical tool for the routine diagnosis and therapeutic evaluation of human schistosomiasis.

Genetic variability among Schistosoma japonicum isolates from different endemic regions in China revealed by sequences of three mitochondrial DNA genes.

The present study examined sequence variation in three mitochondrial DNA (mtDNA) regions, namely cytochrome c oxidase subunit 3 (cox3), NADH dehydrogenase subunits 4 and 5 (nad4 and nad5), among Schistosoma japonicum isolates from different endemic regions in China, and their phylogenetic relationships were re-constructed. A portion of the cox3 gene (pcox3), a portion of the nad4 and nad5 genes (pnad4 and pnad5) were amplified separately from individual trematodes by polymerase chain reaction (PCR) and the amplicons were subjected to direct sequencing. In the mountainous areas, sequence variations between parasites from Yunnan and those from Sichuan were 0.3% for pcox3, 0.0-0.1% for pnad4, and 0.0-0.2% for pnad5. In the lake/marshland areas, sequence variations between male and female parasites among different geographical locations were 0.0-0.3% for pcox3, 0.0-0.7% for pnad4, and 0.0-1.6% for pnad5. Sequence variations between S. japonicum from mountainous areas and those from lake/marshland areas were 0.0-0.5% for pcox3, 0.0-0.7% for pnad4, and 0.0-1.6% for pnad5. Phylogenetic analyses based on the combined sequences of pcox3, pnad4 and pnad5 revealed that S. japonicum isolates from mountainous areas (Yunnan and Sichuan provinces) clustered together. For isolates from the lake/marshland areas, isolates from Anhui and Jiangsu provinces clustered together and was sister to samples from Jiangxi province, while isolates from Hubei and Zhejiang province clustered together. However, isolates from different geographical locations in Hunan province were in different clades. These findings demonstrated the usefulness and attributes of the three mtDNA sequences for population genetic studies of S. japonicum, and have implications for studying population biology, molecular epidemiology, and genetic structure of S. japonicum, as well as for the effective control of schistosomiasis.

Praziquantel and the benzodiazepine Ro 11-3128 do not compete for the same binding sites in schistosomes.

The benzodiazepine Ro 11-3128 (methyl-clonazepam) presents several similarities with praziquantel with regard to its anti-schistosomal mode of action, since both drugs cause spastic paralysis, calcium influx and tegumental disruption in the parasites. In order to know whether the two compounds share the same binding sites in the schistosomes, we performed in vivo and in vitro competition experiments. We took advantage of the fact that Ro 11-3128 is active against immature Schistosoma mansoni (whereas praziquantel is inactive), and praziquantel is active against S. japonicum (which is insensitive to Ro 11-3128). An excess of praziquantel did not inhibit the activity of Ro 11-3128 against immature S. mansoni and an excess of Ro 11-3128 did not inhibit the activity of praziquantel against S. japonicum, suggesting that the schistosome binding sites of the two drugs are different. On the other hand, cytochalasin D, an agent known to perturb--among other things--calcium channel function, was capable of inhibiting the schistosomicidal activity of both praziquantel and Ro 11-3128, thus adding another element of similarity between the two anti-schistosomal agents. A similar, albeit partial, inhibition of the schistosomicidal activity of the two drugs was exerted by some of the classical calcium channel blockers. Taken together, these results suggest that praziquantel and Ro 11-3128, although binding to different schistosome receptor sites, may use the same basic anti-schistosomal effector mechanisms.

Rapid-processing procedure for heat polymerization of polymethyl methacrylate in a pressure cooker with automatic controls.

This study introduces a new rapid-processing procedure for curing polymethyl methacrylate denture base resin in an automatically controlled pressure cooker. The cooker filled with water was inflated with 6 kgf/cm2 air pressure and heated to 120 degrees C (248 degrees F) and maintained for 10 minutes. No significant differences were found between the new pressure cooker method and the conventional method for surface hardness and porosity (p > 0.05). The pressure cooker significantly shortened polymerization time, and the polymerization is controlled automatically.

[Studies on the effect of immunoenhancing reagent "425" on egg granuloma formation in mice infected with Schistosoma japonicum].

Fourteen--34 days after Schistosoma japonicum-infected mice were injected intraperitoneally with immunoenhancing reagent, Chinese angelica root extract ("425"), the specific antibody (IgG) levels were significantly higher in Group III (362.67 +/- 162.21 - 480.00 +/- 289.45) than the control group (66.67 +/- 39.68 - 245.33 +/- 101.56) in experiment one, and in Group II (488.00 +/- 320.38 - 768.00 +/- 267.38) than the control group (256.00 +/- 189.07 - 394.67 +/- 141.06) in experiment two. The egg granulomatous formation were markedly diminished in the injected mice. The ratios of granulomas vs. eggs' mean diameters of Group III of mice were 5.24 +/- 1.04 - 3.95 +/- 0.77 and those of the control group were 6.59 +/- 1.19 - 5.29 +/- 0.94 in experiment one, those of the group II were 3.75 +/- 0.71 - 4.15 +/- 0.73 and those of the control group were 4.94 +/- 0.81 - 5.36 +/- 0.97 in experiment two. Meantime the egg antigen levels in the injected mice might be lower. This study shows that the control of immunomodulation of granulomatous formation in the hosts injected with immunoenhance reagent "425" can be induced.