Evaluation of in vitro antileishmanial efficacy of cyclosporin A and its non-immunosuppressive derivative, dihydrocyclosporin A.

BACKGROUND: New therapeutic drugs are urgently needed against visceral leishmaniasis because current drugs, such as pentavalent antimonials and miltefosine, produce severe side effects and development of resistance. Whether cyclosporine A (CsA) and its derivatives can be used as therapeutic drugs for visceral leishmaniasis has been controversial for many years. METHODS: In this study, we evaluated the efficacy of CsA and its derivative, dihydrocyclosporin A (DHCsA-d), against promastigotes and intracellular amastigotes of Leishmania donovani. Sodium stibogluconate (SSG) was used as a positive control. RESULTS: Our results showed that DHCsA-d was able to inhibit the proliferation of L. donovani promastigotes (IC50: 21.24 µM and 12.14 µM at 24 h and 48 h, respectively) and intracellular amastigotes (IC50: 5.23 µM and 4.84 µM at 24 and 48 h, respectively) in vitro, but CsA treatment increased the number of amastigotes in host cells. Both DHCsA-d and CsA caused several alterations in the morphology and ultrastructure of L. donovani, especially in the mitochondria. However, DHCsA-d showed high cytotoxicity towards cells of the mouse macrophage cell line RAW264.7, with CC50 values of 7.98 µM (24 h) and 6.65 µM (48 h). Moreover, DHCsA-d could increase IL-12, TNF-α and IFN-γ production and decrease the levels of IL-10, IL-4, NO and H2O2 in infected macrophages. On the contrary, CsA decreased IL-12, TNF-α, and IFN-γ production and increased the levels of IL-10, IL-4, NO and H2O2 in infected macrophages. The expression of L. donovani cyclophilin A (LdCyPA) in promastigotes and intracellular amastigotes and the expression of cyclophilin A (CyPA) in RAW 264.7 cells were found to be significantly downregulated in the CsA-treated group compared to those in the untreated group. However, no significant changes in LdCyPA and CyPA levels were found after DHCsA-d or SSG treatment. CONCLUSIONS: Our findings initially resolved the dispute regarding the efficacy of CsA and DHCsA-d for visceral leishmaniasis treatment. CsA showed no significant inhibitory effect on intracellular amastigotes. DHCsA-d significantly inhibited promastigotes and intracellular amastigotes, but it was highly cytotoxic. Therefore, CsA and DHCsA-d are not recommended as antileishmanial drugs.

Direct and Indirect Inhibition Effects of Resveratrol against Toxoplasma gondii Tachyzoites In Vitro.

Toxoplasma gondii is one of the most widespread obligatory parasitic protozoa and infects nearly all warm-blooded animals, leading to toxoplasmosis. The therapeutic drugs currently administered, like the combination of pyrimethamine and sulfadiazine, show high rates of toxic side effects, and drug resistance is encountered in some cases. Resveratrol is a natural plant extract with multiple functions, such as antibacterial, anticancer, and antiparasite activities. In this study, we evaluated the inhibitory effects of resveratrol on tachyzoites of the Toxoplasma gondii RH strain extracellularly and intracellularly. We demonstrate that resveratrol possesses direct antitoxoplasma activity by reducing the population of extracellularly grown tachyzoites, probably by disturbing the redox homeostasis of the parasites. Moreover, resveratrol was also able to release the burden of cellular stress, promote apoptosis, and maintain the autophagic status of macrophages, which turned out to be regulated by intracellular parasites, thereby functioning indirectly in eliminating T. gondii In conclusion, resveratrol has both direct and indirect antitoxoplasma effects against RH tachyzoites and may possess the potential to be further evaluated and employed for toxoplasmosis treatment.

[In vivo and in vitro anti-tumor effect of Sanjie Kangliu Decoction].

OBJECTIVE: To study the anti-tumor effect of Sanjie Kangliu (swelling-dispersing) Decoction in vivo and in vitro. METHODS: The mice bearing tumors (established by subcutaneous inoculation of SRS-82 cells) were fed with Sanjie Kangliu Decoction. The tumor was isolated and weighed to calculate the tumor inhibition rate, followed by morphological observation of the tumor cells under electron microscope. The DNA of the tumor cells was then extracted for agarose gel electrophoresis. In the in vitro experiment, cultured SRS-82 cells were treated with the serum of rats fed with Sanjie Kangliu Decoction, and the cell viability and apoptosis was examined by MTT assay and flow cytometry, respectively. Immunocytochemistry was employed to test the expression intensity of Bcl-2 protein of the rat serum-treated cells, and in situ hybridization performed to detect matrix metalloproteinase (MMP)-2 mRNA expression. RESULTS: In the tumor-bearing mouse models, Sanjie Kangliu Decoction administration resulted in significant reduction of the tumor mass and apoptosis of the tumor cells as shown by typical apoptotic changes of the cell morphology and presence of DNA ladder in agarose gel electrophoresis. In comparison with the mice without the decoction treatment, the treated mice showed significantly lower tumor cell viability, higher cell apoptosis rate, and weaker Bcl-2 expression. The expression of MMP-2 mRNA showed no distinct difference between the groups. CONCLUSION: Sanjie Kangliu Decoction has obvious anti-tumor effect both in vivo and in vitro possibly through the mechanism of inducing tumor cell apoptosis by reducing Bcl-2 expression without affecting the expression of MMP-2 mRNA.

[Inhibitory effect of Biejiajianwan pills on tumor growth and proliferating cell nuclear antigen expression in mice bearing H22 cell-induced tumor].

OBJECTIVE: To study the anti-tumor effect of Biejiajianwan pills and explore its mechanism. METHODS: Mice bearing neoplasm induced by H22 cell inoculation were randomized into 4 groups, namely the negative control group, positive control group (with cyclophosphamide treatment), and high- and low-dose Biejiajianwan pill group. The tumor-bearing mice received corresponding treatments for 15 consecutive days, after which their thymuses and spleens were isolated and weighed to calculate the thymus and spleen indices. The tumor mass was also weighed and the tumor inhibition rate calculated. Immunohistochemistry was performed to evaluate the expression intensity of proliferating cell nuclear antigen (PCNA) in the tumor tissue. RESULTS: The pills significantly inhibited the growth of the implanted tumor and reduced the expression of PCNA in the tumor. CONCLUSION: One of the possible mechanisms of the anti-tumor effects of Biejiajian pills might be associated with the inhibition of PCNA expression in the tumor.