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Objective: To investigate the effects of modification of acellular bovine pericardium with 1-ethyl-3-(3- dinethylami-nopropyl) carbodimide (EDC)/N-hydroxysuccininide (NHS) or genipin and find out the best crosslinking reagent., Methods: The cellular components of the bovine pericardiums were removed. The effects of decellularization were tested by HE staining. The acellular bovine pericardiums were crosslinked with EDC/NHS (EDC/NHS group) or genipin (genipin group). The properties of the crosslinked acellular matrix were evaluated by scanning electron microscope (SEM), matrix thickness, crosslinking index, mechanical property, denaturation temperature, enzymatic degradation, and cytotoxicity test before and after the crosslinking. Acellular bovine pericardium (ABP group) or normal bovine pericardium (control group) were harvested as controls., Results: SEM showed that collagen fibers were reticulated in bovine pericardial tissues after crosslinked by EDC/NHS or genipin, and relative aperture of the collagen fiber was from 10 to 20 μm. The thickness and denaturation temperature of the scaffolds were increased significantly after crosslinking with EDC/NHS or genipin (P < 0.05), while there was no significant difference between EDC/NHS group and genipin group (P > 0.05). The difference had no statistical significance in crosslinking index between EDC/NHS group and genipin group (t = 0.205, P = 0.218). The degradation rate in EDC/NHS group and genipin group was significantly lower than that in ABP group and control group (P < 0.05). Elastic modulus and fracture stress in EDC/NHS group and genipin group were significantly lower than those in ABP group (P < 0.05), but there was no significant difference among EDC/NHS group, genipin group, and control group (P > 0.05). The break elongation in EDC/NHS group and genipin group were significantly increased than those in ABP group and control group (P < 0.05). The difference had no statistical significance in stability and mechanical properties between EDC/NHS group and genipin group (P > 0.05). Cytotoxicity of genipin crosslinked tissue (grade 1) were much lower than that of EDC/NHS (grade 2) at 5 days., Conclusion: Acellular bovine pericardium crosslinked with genipin has better biocompatibility than EDC/NHS.

Objective: To observe the descending modulation of cardiac nociception by the rostral ventromedial medulla (RVM) in rats., Methods: A rat model of cardiosomatic motor reflex (CMR) was established by injecting capsaicin into the pericardial sac to induce cardiac nociception, and the electromyogram (EMG) response of the dorsal spinotrapezius muscle was studied. The RVM was electrically stimulated (25, 75 and 100 µA) or destroyed to examine whether RVM exerted descending modulation on cardiac nociception., Results: Electrical stimulation of the RVM at 8 sites produced intensity-dependent inhibition of EMG responses to noxious cardiac stimulus (F[2,21]=43.188, P=0.001). Electrical stimulation at 3 sites caused facilitated EMG responses, but the increased magnitude of the EMG was not dependent on stimulation intensity (F[2,6]=0.884, P=0.461). Stimulation at 11 sites produced biphasic effects: at a low intensity (25 µA), the elicited EMG magnitude was significantly larger than baseline (P<0.05), and at greater intensities (75/100 µA), the stimulation caused suppression of the EMG magnitude to a level significantly lower than the baseline (P<0.05). Electrolytic lesion of the RVM resulted in significantly increased EMG responses compared with the baseline and sham lesion group., Conclusion: Cardiac nociception evoked by capsaicin stimulation is subjected to descending biphasic modulation by the RVM, which produces predominantly descending inhibition on heart pain.

Objective: To evaluate the hemocompatibility of glutaraldehyde (GA)-tanned bovine pericardium additionally treated by sodium bisulfite (SOB) solution., Methods: The hemocompatibility of GA-tanned bovine pericardium treated by SOB solution is evaluated by using dynamic clotting time test, blood platelet adhension test, D-dimeride determination, and complement activation test. The GA-tanned bovine pericardium was used as control., Results: The curve of absorbance-clotting time of two kinds of bovine pericardium was similar in dynamic clotting time test. There was no significant difference between SOB-treated and control groups in blood platelet adhension test. The D-dimeride contents of all bioprostheses were at normal level, and the D-dimeride content of GA-tanned bovine pericardium treated by SOB solution was significantly lower than that of control group (P < 0.05). In complement activation test, the level of complement C3a in SOB-treated group was significantly lower than that in control group (P < 0.05)., Conclusion: GA-tanned bovine pericardium treated by SOB solution meets the demands of cardiac interstitial implanted materials in hemocompatibility.

To observe the growth of rat bone marrow mesenchymal cells (BMMCs) on decellular bovine pericardia in vitro and to investigate the effect of proteins pre-coating on cells retention and proliferation, bovine pericardia were decellularized using trypsin, DNase and Triton X-100 respectively. Then three proteins (fibronetin (FN), gelatin, collagen I) were coated on the surfaces of the bovine pericardia separately. BMMCs were harvested from rat thighbone marrow , then expanded and seeded onto decellular bovine pericardia with the proteins pre-coated . Decelluar bovine pericardia without coating were used as controls. The retention and growth of BMMSCs were observed by Hochest staining and analyzed by MTT method. It was shown that the retention and proliferation of BMMCs on FN group and gelatin group were significantly enhanced comparing with those on collagen I group and control group (P < 0.001). There was no significant difference between FN group and gelatin group (P > 0.05), nor between collagen I group and control group (P > 0.05). We conclude that the retention and proliferation of seeding cells on FN and gelatin could be significantly improved on decellular bovine pericardia (DBP) but not on collagen I.

Bovine pericardium is mainly composed of collagen which has many good properties of extracellular matrix. This in vitro study was performed to quantitatively investigate and compare the degradation behavior of the bovine pericardial, which were modified with different methods. The purpose is to find out one method that presents not only the degradation regularity of material but also minimum antigenicity, so that bovine pericardium can be used as a well degradable guide tissue regeneration material. The results of the experiments showed that the bovine pericardium treated with ethanol is more appreciated than that treated with epoxy cross-linking agent or glutaraldehyde. The mensuration of protein and special amino acid may be a useful base of constructing a mathematic model for further researches on the rate of degradation of bovine pericardium.

Objective: To investigate a method of autologous pericardial heart valves (APHV) treatment beneficial to endothelialization., Methods: Bovine aortic endothelium cells (BAEC) were cultured and seeded in autologous pericardial pieces treated with different methods in vitro. The influence of different treatment on cellular growing, amount of 3H-TdR incorporation and PGI2 releasing, contents of intracellular cAMP was studied by means of transmission electron microscopy, 3H-TdR incorporation experimentation and radioimmunity method., Results: The BAEC seeded in bovine pericardial pieces treated with three different concentrations of GA 10 minutes could alive. In the group with method of treatment with 0.2% GA, the growing of BAEC was best, the cellular density and the amount of 3H-TdR incorporation at different time spots were higher than that of the other groups and it did not enhance the abnormal synthesis of cAMP and PGI2 in BAEC., Conclusion: The pericardium treated with 0.2% GA 10 minutes has little toxicity to endothelium cell, which indicates that this method is beneficial to endothelialization of APHV postoperatively.

Objective: To observe the regulating effects of vascular endothelial growth factor (VEGF) and angiotensin II (ANG II) on the frog's pericardium, lymphatic stomata, and angionesis so as to understand their influences on the mesothelial permeability, lymphatic stoma regulation, and myocardial hypertrophy., Methods: VEGF and ANG II were injected into the frog's peritoneal cavity. The results were obtained by using LM, SEM, and computerized imaging analysis., Results: Scattered distributed pericardial stomata were found on the parietal pericardium of the frog with a few sinusoid mesothelial cells. Since there were no blood vessels in the frog's myocardium, its blood supply was directly from the cardiac chambers flowing into the trabecular spaces of the myocardium. The average diameters of the pericardial stomata in VEGF and ANG II groups were 1.50 and 1.79 microns respectively, which were much larger than those in the control group (0.72 micron, P < 0.01). The average distribution density of the stomata was 8.25/0.1 mm2 and 12.80/0.1 mm2 in VEGF and ANG II groups, which was also much higher than that in the control group (3.57/0.1 mm2, P < 0.01). The sinusoid areas in VEGF and ANG II groups were 2443.95 microns 2/0.1 mm2 and 2121.79 microns 2/0.1 mm2, which was larger than that in the control group (995.08 microns 2/0.1 mm2, P < 0.01). No angionesis were found in the frogs of the experimental groups., Conclusions: VEGF and ANG II strongly regulate the pericardial stomata by increasing their numbers and openings with larger diameters and higher distribution density. They can also increase the sinusoid areas with the result of the higher permeability of the pericardium, which clealy indicated that VEGF and ANG II can speed up the material transfer of the pericardial cavity and play an important role in preventing myocardial interstitial edema. Yet there is no strong evidence showing angionesis in the myocardium.