Ingrid Samantha Tavares de Figueiredo

Involvement of NO in the inhibitory effect of Calotropis procera latex protein fractions on leukocyte rolling, adhesion and infiltration in rat peritonitis model

AIM OF THE STUDY The latex of Calotropis procera has been used in the traditional medicinal system for the treatment of leprosy, ulcers, tumors, piles and diseases of liver, spleen, abdomen and toothache. It comprises of a non-dialyzable protein fraction (LP) that exhibits anti-inflammatory properties and a dialyzable fraction (DF) exhibiting pro-inflammatory properties. The present study was carried out to evaluate the effect of LP sub-fractions on neutrophil functions and nociception in rodent models and to elucidate the mediatory role of nitric oxide (NO). MATERIAL AND METHODS The LP was subjected to ion exchange chromatography and the effect of its three sub-fractions (LP(PI), LP(PII) and LP(PIII)) thus obtained was evaluated on leukocyte functions in the rat peritonitis model and on nociception in the mouse model. RESULTS LP sub-fractions exhibit distinct protein profile and produce a significant decrease in the carrageenan and DF induced neutrophil influx and exhibit anti-nociceptive property. The LP and its sub-fractions produced a marked reduction in the number of rolling and adherent leukocytes in the mesenteric microvasculature as revealed by intravital microscopy. The anti-inflammatory effect of LP(PI), the most potent anti-inflammatory fraction of LP, was accompanied by an increase in the serum levels of NO. Further, our study shows that NO is also involved in the inhibitory effect of LP(PI) on neutrophil influx. CONCLUSIONS Our study shows that LP fraction of Calotropis procera comprises of three distinct sets of proteins exhibiting anti-inflammatory and anti-nociceptive properties of which LP(PI) was most potent in inhibiting neutrophil functions and its effects are mediated through NO production.

Proteins derived from latex of C. procera maintain coagulation homeostasis in septic mice and exhibit thrombin- and plasmin-like activities

The proteins derived from the latex (LP) of Calotropis procera are well known for their anti-inflammatory property. In view of their protective effect reported in the sepsis model, they were evaluated for their efficacy in maintaining coagulation homeostasis in sepsis. Intraperitoneal injection of LP markedly reduced the procoagulation and thrombocytopenia observed in mice infected with Salmonella; while in normal mice, LP produced a procoagulant effect. In order to understand its mechanism of action, the LP was subjected to ion-exchange chromatography, and the three subfractions (LPPI, LPPII, and LPPIII) thus obtained were tested for their proteolytic effect and thrombin- and plasmin-like activities in vitro. Of the three subfractions tested, LPPII and LPPIII exhibited proteolytic effect on azocasein and exhibited procoagulant effect on human plasma in a concentration-dependent manner. Like trypsin and plasmin, these subfractions produced both fibrinogenolytic and fibrinolytic effects that were mediated through the hydrolysis of the Aα, Bβ, and γ chains of fibrinogen and α-polymer and γ-dimer of fibrin clot, respectively. This study shows that the cysteine proteases present in the latex of C. procera exhibit thrombin- and plasmin-like activities and suggests that these proteins have therapeutic potential in various conditions associated with coagulation abnormalities.

Preclinical anticancer effectiveness of a fraction from Casearia sylvestris and its component Casearin X : in vivo and ex vivo methods and microscopy examinations

ETHNOPHARMACOLOGICAL RELEVANCE Casearia sylvestris (Salicaceae) is found in South America and presents antiulcerogenic, cytotoxic, antimicrobial, anti-inflammatory and antihypertensive activities. AIM OF THE STUDY To assess the in vivo and ex vivo antitumor action of a fraction with casearins (FC) and its main component - Casearin X-isolated from C. sylvestris leaves. MATERIALS AND METHODS Firstly, Sarcoma 180 bearing Swiss mice were treated with FC and Cas X for 7 days. Secondly, BALB/c nude animals received hollow fibers with colon carcinoma (HCT-116) or glioblastoma (SF-295) cells and were treated with FC for 4 days. On 5th day, proliferation was determined by MTT assay. RESULTS FC 10 and 25mg/kg/day i.p. and 50mg/kg/day oral and Cas X 25mg/kg/day i.p. and 50mg/kg/day oral revealed tumor growth inhibition rates of 35.8, 86.2, 53.7, 90.0 and 65.5% and such tumors demonstrated rare mitoses and coagulation necrosis areas. Similarly, FC reduced multiplying of HCT-116 and SF-295 cells when evaluated by the Hollow Fiber Assay (2.5 and 5mg/kg/day i.p. and 25 and 50mg/kg/day oral), with cell growth inhibition rates ranging from 33.3 to 67.4% (p<0.05). Flow cytometry experiments revealed that FC reduced membrane integrity and induced DNA fragmentation and mitochondrial depolarization (p<0.05). CONCLUSIONS FC and Cas X were efficient antitumor substances against murine and human cancer cells and caused reversible morphological changes in liver, kidneys and spleens, emphasizing clerodane diterpenes as an emerging class of anticancer molecules.

Vascular permeability, neutrophil migration and edematogenic effects induced by the latex of Cryptostegia grandiflora.

Inflammatory responses have been described as occurring after exposure to some latex materials. In this study pro-inflammatory activity in the latex of Cryptostegia grandiflora was investigated. The soluble proteins of the latex (CgLP) were isolated from the whole latex and evaluated by in vivo assays. CgLP induced strong inflammatory activity mediated by neutrophil migration, enlarging vascular permeability and increasing myeloperoxidase activity locally in rats. CgLP-induced inflammation was observed in peritonitis, paw edema and air push models. In addition, CgLP caused hyperemia in a healing model. The peritonitis effect was lost when CgLP was previously boiled suggesting the involvement of pro-inflammatory proteins. Thioglycollate increased the neutrophil migration induced by CgLP, but not by fMLP. Mast cell depletion provoked by 40/80 compound did not modify the course of inflammation triggered by CgLP, being similar to fMLP, which suggested that neutrophil migration was induced by direct mechanism mediated by macrophages. Neutrophil migration stimulated by CgLP was strongly inhibited by Dexamethasone and to a lesser extent by Thalidomide, indicating the involvement of cytokines in mediating neutrophil infiltration. Celecoxib and Indomethacin were inhibitory suggesting the involvement of prostaglandins. Cimetidine was effective only in the initial phase of edema. PCA 4248 was ineffective. It is concluded that the latex of C. grandiflora is a potent inflammatory fluid, and also that laticifer proteins may be implicated in this process.

Side-Effects of Irinotecan (CPT-11), the Clinically Used Drug for Colon Cancer Therapy, Are Eliminated in Experimental Animals Treated with Latex Proteins from Calotropis procera (Apocynaceae).

Intestinal mucositis (IM) is the critical side effect of irinotecan (CPT‐11), which is the front‐line drug used for the treatment of colorectal cancer. This study aimed to evaluate the effectiveness of latex proteins (LP) from Calotropis procera to prevent IM and diarrhea in animals. Swiss mice were treated daily with saline or LP (1, 5, or 50 mg/kg, i.v.) 24 h prior to CTP‐11 (75 mg/kg/4 days, i.p) and for additional 6 days. Animal survival, body weight variation, and diarrhea were registered. After animal sacrifice (day 7 post first injection of CPT‐11), intestinal samples were collected to study morphology and inflammatory parameters. Animals given LP exhibited improved parameters (survival, body weight, and absence of diarrhea) as compared with the CPT‐11 control. The severity of IM observed in animals given CPT‐11 was reduced in animals treated with LP. Treatment with LP also prevented the reduction in the villus/crypt ratio promoted by CPT‐11. The rise in MPO activity and pro‐inflammatory cytokines, over‐contractility of the smooth muscle, and diarrhea were all abrogated in LP‐treated mice. Markedly reduced immunostaining intensity for COX‐2, TNF‐α, IL‐1β, iNOS, and NF‐κB was observed in the intestinal tissue of animals treated with LP. The side‐effects of CPT‐11 were eliminated by LP treatment in experimental animals and improved clinical parameters characteristic of IM All known biochemical pathogenesis. Copyright

The Preventive Effect on Ethanol-Induced Gastric Lesions of the Medicinal Plant Plumeria rubra: Involvement of the Latex Proteins in the NO/cGMP/KATP Signaling Pathway

Plumeria rubra (Apocynaceae) is frequently used in folk medicine for the treatment of gastrointestinal disorders, hepatitis, and tracheitis, among other infirmities. The aim of this study was to investigate the gastroprotective potential of a protein fraction isolated from the latex of Plumeria rubra (PrLP) against ethanol-induced gastric lesions and describe the underlying mechanisms. In a dose-dependent manner, the pretreatment with PrLP prevented ethanol-induced gastric lesions in mice after single intravenous administration. The gastroprotective mechanism of PrLP was associated with the involvement of prostaglandins and balance of oxidant/antioxidant factors. Secondarily, the NO/cGMP/KATP pathway and activation of capsaicin-sensitive primary afferents were also demonstrated as part of the mechanism. This study shows that proteins extracted from the latex of P. rubra prevent gastric lesions induced in experimental animals. Also, the results support the use of the plant in folk medicine.