Nitric Oxide Regulates The Lymphatic Reactivity Following Hemorrhagic Shock Through Atp-Sensitive Potassium Channel.

Lymphatic reactivity has been shown to exhibit a biphasic change following hemorrhagic shock, and nitric oxide (NO) is involved in this process. However, the precise mechanism responsible for NO regulation of the lymphatic reactivity along with the progression of hemorrhagic shock is unclear. Therefore, the present study was to investigate how NO participates in regulating the shock-induced biphasic changes in lymphatic reactivity and its underlying mechanisms. First, the expressions or contents of inducible NO synthase, nitrite plus nitrate, and elements of cAMP-PKA-KATP and cGMP-PKG-KATP pathway in thoracic ducts tissue were assessed. The results revealed that levels of nitrite plus nitrate, cAMP, cyclic guanosine monophosphate (cGMP), p-PKA, and p-PKG were increased gradually along with the process of shock. Second, the roles of cAMP-PKA-KATP and cGMP-PKG-KATP in NO regulating lymphatic response to gradient substance P were evaluated with an isolated lymphatic perfusion system. The results showed that the NOS substrate (L-Arg), PKA donor (8-Br-cAMP) decreased the reactivity of shock 0.5 h-lymphatics, and that the PKA inhibitor (H-89) and KATP inhibitor (glibenclamide) restrained the effects of L-Arg while glibenclamide abolished the effects of 8-Br-cAMP. Meanwhile, NOS antagonist (L-NAME), protein kinase G (PKG) inhibitor (KT-5823), and soluble guanylate cyclase inhibitor (ODQ) increased the reactivity of shock 2 h-lymphatics, whereas KATP opener (pinacidil) inhibited these elevated effects induced by either L-NAME, ODQ, or KT-5823. Taken together, these results indicate that NO regulation of lymphatic reactivity during shock involves both cAMP-PKA-KATP and cGMP-PKG-KATP pathways. These findings have potential significance for the treatment of hemorrhagic shock through regulating lymphatic reactivity.

[Reactivity to substance P of isolated lymphatics in hemorrhagic shock rat].

OBJECTIVE: To observe the change of lymphatic reactivity to substance P (SP) during the process of hemorrhagic shock (HS) with a technique of lymphatic perfusion in vitro in this study. METHODS: Male Wistar rats were randomly divided into control group (surgical procedure only) and HS group (the rats in this group were further divided into five subgroups: shock 0 h, 0.5 h, 1 h, 2 h and 3 h groups after duplicating the HS model with method of bloodletting to mean arterial blood pressure was 40 mmHg through the femoral venous). Thoracic ducts were separated from HS rats at the corresponding time points in each group. A segment of thoracic duct was pressed and perfused in vitro at transmural pressure of 3 cm H2O, and then stimulated with gradient SP respectively. The end systolic diameter, end diastolic diameter, contraction frequency (CF) and passive diameter of isolated lymphatics were measured, while the contraction amplitude (CA), tonic index (TI) and fractional pump flow (FPF) were calculated, and the different values between pre- and post- administration of SP of CF, CA, TI and FPF were calculated and expressed as Delta CF, Delta TI, Delta CA and Delta FPF to further assess the reactivity of lymphatics. RESULTS: After SP incubation, the Delta CF, Delta TI, Delta CA and Delta FPF of 0 h- and 0.5 h shocked lymphatics were significantly increased when compared with that of control group on one or several concentrations. The Delta CF (at 3 x 10(-7) mol/L of SP) and Delta TI (1 x 10(-7) mol/L) of 2 h- shocked lymphatics and the Delta CF (1 x 10(-7) mol/L, 3 x 10(-7) mol/L), Delta TI (1 x 10(-7) mol/L) and Delta CA (1 x 10(-7) mol/L) of 3 h- shocked lymphatics were all significantly reduced when compared with control group. CONCLUSION: The reactivity of lymphatics to SP presented a biphasic change during the process of HS: increase in early phase and decline in later stage.

[Nitric oxide modulates biphasic changes of isolated lymphatic contraction in hemorrhagic shock rats].

The aim of the present study was to investigate the changes of lymphatic contraction after hemorrhagic shock in vitro and the underlying role of nitric oxide (NO). Rat thoracic duct segments were isolated at 0, 0.5, 1, 2 and 3 h after hemorrhagic shock. Using Pressure Myograph System, we determined contraction frequency (CF), end systolic diameter (ESD), end diastolic diameter (EDD) and passive diameter (PD) of isolated rat lymphatics under different transmural pressures (1, 3, 5, 7 and 9 cmH(2)O), then calculated contraction amplitude (CA), tonic index (TI) and fractional pump flow (FPF) of lymphatics. The results showed that in several transmural pressures, lymphatic CF, TI and FPF were significantly higher in shock 0 h and shock 0.5 h groups than those in control group (sham operation group). With the development of shock, lymphatic CF, TI and FPF decreased significantly in shock 2 h and shock 3 h groups compared with those in control group. We further discovered the role of NO in the changes of lymphatic contraction after hemorrhagic shock. Under 3 cmH(2)O transmural pressure, the changes of lymphatic contraction in shock 0.5 h and shock 2 h groups were analyzed following the incubation with several NO-related drugs alone or in combination. And the results showed that NO donor L-Arg reduced CF, TI and FPF in shock 0.5 h group to the control levels, while soluble guanylate cyclase inhibitor ODQ suppressed the effect of L-Arg. Moreover, NOS inhibitor L-NAME elevated the CF, TI and FPF of 2 h shock lymphatics to the control levels, while phosphodiesterase inhibitor aminophylline (AP) suppressed the effect of L-NAME. These results suggest that the lymphatic contractile activity exhibits a biphasic change during hemorrhagic shock, increasing in early phase and declining in later stage. And NO plays a major regulating role in the biphasic change of lymphatic contraction in hemorrhagic shock rats via cGMP pathway.

Detection of H. pylori antibody profile in serum by protein array.

AIM: To detect multiple H. pylori antibodies in serum samples of individuals who carry H. pylori by protein array. METHODS: Recombinant H. pylori antigens, urease B subunit (UreB), vacuolating toxin A (VacA) and cytotoxin associated gene A protein (CagA), were prepared and immobilized in matrixes on nitrocellulose membrane by robotics to bind the specific immunoglobulin G (IgG) antibodies in serum. Staphylococcus protein A (SPA) labeled by colloid gold was used to integrate the immuno-complex and gave red color signal. The scanner based on charge-coupled device (CCD) could collect the image signal and convert it into digital signal. RESULTS: When human IgG was printed on the membrane in increasing concentrations and incubated with immunogold, a linear dose response curve was obtained and the detection limit for IgG was about 0.025 ng. The cutoff values, which were defined as the mean grey level plus 3 times of standard deviation, were 27.183, 28.546 and 27.402, for anti-UreB IgG, anti-CagA IgG and anti-VacA IgG, respectively, as 400 human serum samples with negative H. pylori antibodies were detected by the protein array. When 180 serum samples from patients in hospital were employed for detection of IgG against UreB, CagA and VacA, the sensitivity of the protein array was 93.4%, 95.4%, 96.0%, and the specificity was 94.8%, 94.4% and 97.5%, respectively, as compared with the results obtained by ELISA. The assay also showed high reproducibility, uniformity and stability, and the results were available within 30 min. CONCLUSION: The protein array is a very practical method for rapid detection of multiple antibodies in serum samples. It is especially useful for large scale epidemiological investigation of the infection of H. pylori.