Effect Of Oleum Cinnamomi On Mice Viral Myocarditis

As an increasing disease incidence of viral myocarditis (VMC) for the past few years, and there has been no therapeutics with specific effect for VMC up to now, the research and development of a medicine in force to to treat VMC becomes a hot issue attracting attention. In the past, to appraise curative effect of drug on VMC, people adopt the method of drug-administering immediately after establishing the VMC model of mice, however, this method only shows the drug has an prophylaxis effect on VMC and did not evaluate the drug has an treatment effect on VMC . Oleum Cinnamomi (OC) is official drug included in the Pharmacopeia , the Pharmacopeia regulates the content of cinnamic aldehyde (CA) in OC shoud be more than 75%. CA has simple chemcial constitution and can be synthesis by artificial, it has lower price than OC.Aim: With the method of drug-administering in 72 hours after establish the VMC model of mice, the research evaluates the treatment effect of OC and CA on the mice VMC model, and presume active component and. mechanism of action of OC treatment effect on VMC.These provids a pharmacodynamics basis on which to research and develop a new medicine.Methods:1. Investigate of LD50 of CA and OC: Fifty Kunming mice were randomLy divided into 5 groups. Percentage of female and male is 50% in each group. Five groups were treated with 2660mg/k,2394mg/kg,2155mg/kg,1939 mg/kg ,1745 mg/kg CA which diluted by tween-80. In following seven days,succession observation and record toxic reaction and death condition of animals was executed. Calculate LD50 of CA by statistically method. LD50 of OC was evaluated in same way.2. Duplicate the VMC model: Thirty male BALB/c mice were randomly divided into A,B,C three groups and were inoculated with Virus titer is 0.54×10⁹ ,1.09×10¹0,2.18×10¹0 pfu/L CVB₃ 0.1mL i.p. To confirm whether or not the mode doing well, on the 5st day after viral inoculation, two mice were randomly take from each groups to detect Coxsackievirus IgM in serum and heart histopathologic examination. To investigate the best does of virus inoculation, on the 21st day after viral inoculation, the death rate was calculated, then the survival mice were killed and myocardium histopathologic examination was made.3. The pharmacodynamics experiment of the therapeutic effect of OC on mice VMC: One hundred and sixty six male BALB/c mice were randomLy divided into 6 groups. Group control (n= 16) was treated with NS i.p., Other five groups (n=30 in each groups) were inoculated O.lmL CVB3 i.p. and bred in insulation lab. 72h after viral inoculated, Groups OC were respectively administered 49.1, 36.7, 26.5 mg/kg OC; Model group was treated with normal saline(NS) in same way; Group RA was treated with 50 mg/kg Radix Astragali Injective Solution i.p. as positive control. All groups were given drugs for one week. On the 10th day after after viral inoculation, the heart histopathologic changes, the content of CK, CK-MB and LDH in mice blood serum and MAD, SOD in myocardium were observed. On the 21st day after after viral inoculation, The death rate , myocardium histopathologic examination were made.4. The pharmacodynamics experiment of the therapeutic effect of CA on miceVMC: 174 male BALB/c mice were randomLy divided into 6 groups. The control group (n=24) was treated with normal saline i.p. Other five groups (n=30 in each group) were inoculated with CVB3 i.p. and breded in a lab insulated from the control group. The drug was administered at 72h after viral inoculation: Groups CA (37.5, 28.1, 22.5 mg/kgCA, i.g); RA groups (50 mg/kg RA i.p.); model group and normal control group (2500 mg/kgNS i.g). The drug was administered continuously for 7 days. After viral inoculation 7th day, a pathologic examination was made and the content of CK, CK-MB, LDH were examined; on the 14th day after d viral inoculation, the content of NO was examined in mice blood serum , the content of MAD and SOD were detected in mice myocardium tissue homogenate, immunohistochemistry detect of iNOS and the microscopic examination were made in mice myocardium tissue. On the 21st day, the myocardium underwent the microscopic examination. Results:1. Investigate of LD₅0 of CA and OC: LD₅0 of CA is 2350mg/kg; LD₅0 of OC is 2816mg/kg.2. Duplicate the VMC model: On the 5st day after viral inoculation, the results of detect Coxsackievirus IgM in mice serum in each group show masccline and myocardium histopathologic examination show inflammatory infiltration caused by a great quantity lymphocyte .On the 21st day after viral inoculation, the death rate of A,B,Cgroups were 100%,75%, 50% respectivlly and the histopathologic changes (necrosis, degeneration and cellular infiltration) in C group were lighter in B group.3. The phannacodynamics experiment of the therapeutic effect of OC on mice VMC: In 49.7 and 36.7 mg/kg OC groups, rusults show that thehistopathologic Score , the content of CK,CK-MB in mice blood serum and MAD in myocardium were lower, and the activities of SOD was higher than in Model group(P<0.05).The death rate was lower(P<0.05),the histopathologic changes (necrosis, degeneration and cellular infiltration) were lighter in OC groups than in Model group at 21th day after viral inoculation. 4. The pharmacodynamics experiment of the therapeutic effect of CA on mice VMC: In 37.5, 28.1, and 22.5 mg/kg CA groups, it shows that the release of the content of CK, CK-MB and LDH at the subac stage (on the 7th day after viral inoculation) could be reduced (P<0.05). The content of NO in the blood serum was reduced (P<0.01); the content of MAD and iNOS in the serum in myocardium were lowered, the necrosis and calcification of the mice myocardium at the subacute stage(on the 11th day after drug—administered) were mitigated. Compared with the model group, the difference in pathological scores was remarkable (P<0.05). However, in the 22.5mg/kg CA treatment group, it shows that the inflammatory reaction of myocardium at the chronic stage (on the 18th day after drug—administered)could be abated. Conclusion:1. LD50 of OC is 2816mg/kg . LD50 of CA is 2350mg/kg;2. The achievement ratio of reproduct the VMC model by 0.54×10, 1.09×10¹0,2.18×10¹0 pfu/L CVB3 i.p 0.1mL is 100%. The results of the death rate and the histopathologic examination show 1.09×10¹0 pfu/L CVB3 i.p O.lmL is the best does .The method of drug-administering in 72 hours after establishing the VMC model choud appraise the drug treatment effect on VMC.3. CA has same significant therapeutic effect with OC on CVB3 induced mice VMC. We presume CA maybe the object component of OC treat VMC. Its acting mechanism may be related to the selective inhibition of iNOS and thus immunity adjustment.