This work aimed to evaluate the activity of a lipid transfer protein isolated from Morinda citrifolia L. seeds, McLTP 1 , on the development of intestinal mucositis following irinotecan administration. McLTP 1 (0.5, 2 and 8 mg/kg, i.v.) was injected into mice 1h before irinotecan administration (75 mg/kg, i.p.; 4 days), and then for additional 6 days. Seven days after the rst dose of irinotecan, diarrhea was assessed and the intestine was removed for histological evaluation, assessment of intestinal over-contractility, measurement of myeloperoxidase (MPO), glutathione (GSH), malondialdehyde (MDA), proin ammatory cytokines and chemokine (IL-1, IL-6, and KC levels -a murine homolog of human IL-8 chemokine), analysis of cyclooxygenase 2 (COX-2), nuclear factor kappa B (NF-κB) and nitric oxide synthase (iNOS) expression. At the two highest doses, McLTP 1 administration decreased mortality and diarrhea. McLTP 1 (8 mg/kg, i.v.) signi cantly prevented irinotecan-induced intestinal damage and led to a reduction in overcontractility of the intestinal muscle (p < 0.05). Moreover, McLTP 1 decreased the MPO, IL-1β, IL-6, and KC levels signi cantly by 74.7%, 42%, 92.9%, and 95.9%, respectively. Also, the expression of COX-2, NF-κB, and iNOS was reduced. Pretreatment with McLTP 1 (8mg/kg; i.v.) signi cantly reduced the MDA level and increased the duodenum homogenates' GSH level. Our study provides a potential new therapeutic for preventing irinotecan-induced mucositis, improved clinical parameters, reduced in ammation, and oxidative damage.

Hemorrhagic cystitis is a side effect of chemotherapy induced by an antineoplastic agent from the oxazaphosphorine group (ifosfamide and cyclophosphamide), resulting from the formation of the urotoxic metabolite acrolein. Morinda citrifolia Linn., popularly known as noni, is a species of Rubiaceae, where it is used from the root to the fruit for therapeutic purposes. From the seeds, a thermostable protein called McLTP1 (9.4 kDa) was extracted, among its therapeutic effects, it showed anti-inflammatory, gastroprotective, antibacterial and antinociceptive activity. Thus, the objective of this study is to evaluate the protective effect and the possible mechanism of action of a protein isolated from the seed of Morinda citrifolia (McLTP1) in hemorrhagic cystitis induced by ifosfamide in mice. Hemorrhagic cystitis was induced by intraperitoneal (i.p) administration of ifosfamide (IFO) in a single dose of 400mg/kg, according to a standardized protocol, in male balb/c mice. The experimental group treated with the uroprotective drug, mesna (80 mg/kg; i.p), received a pretreatment 30 minutes before, 4 and 8 hours after IFO. Treatment with McLTP1 was divided into two protocols, the first to define the best dose through a dose-response curve, where a pre-treatment was performed three days before cystitis induction, with McLTP1 administered at doses of 10, 20 or 40mg/kg (i.p), and two treatments 2 and 4 hours after IFO administration, evaluating its effect on bladder wet weight, edema and hemorrhage scores, and neutrophilic infiltrate. In the second protocol, only the best dose was used for the analysis of its effect on the hemorrhagic cystitis model. After 12 hours of hemorrhagic cystitis induction, the animals were euthanized by a high anesthetic dose. Subsequently, the bladders were removed, weighed and kept in 10 per cent buffered formalin for histological, immunohistochemical (COX-2 and TNF-a), immunofluorescence (NF-kB and F4-80) analyses, or stored at -80C for of MPO, vascular permeability, hemoblobin, cytokines (TNF-a;, IL-1B;, IL-6, IL-10, IL-4, IL-33), enzymes (iNOS and COX-2) and markers of oxidative stress (MDA, NO, GSH, SOD and CAT). The adopted experimental procedures were approved by the Animal Research Ethics Committee through protocol number 23170920-0. Treatment with McLTP1 reduced bladder wet weight at the three respective doses mentioned above, however, it was observed the reduction of toxicity parameters (macroscopic edema and hemorrhage scores) only at the lowest dose (10 mg/kg), as well as MPO activity at doses of 10 and 20 mg/kg (p<0.05). results, the lowest dose was chosen for subsequent results. McLTP1 (10 mg/kg) was able to promote permeability reduction and vascular and hemoglobin in the bladder through quantification by the evans blue method and cyanmethemoglobin, respectively (p<0.05). In addition, it had a protective effect by attenuating inflammatory scores and preserving the structure of the urothelium. The anti-inflammatory activity was demonstrated through the significant decrease of the cytokines TNF-a; IL-B;, IL-6 and increase of IL-10; reduced expression of COX-2, NF-kB and F4/80, and gene expression of IL-33, IL-4 and iNOS (p<0.05). McLTP1 also showed antioxidant activity, being able to reduce MDA and NO and increase levels of GSH, SOD and CAT (p<0.05). From the presented data, we can infer that McLTP1 is a potential uroprotector in the prevention of ifosfamide-induced hemorrhagic cystitis in mice by reducing inflammatory parameters and antioxidant activity.

This work aimed to evaluate the activity of a lipid transfer protein isolated from Morinda citrifolia L. seeds, McLTP1, on the development of intestinal mucositis following irinotecan administration. McLTP1 (0.5, 2 and 8 mg/kg, i.v.) was injected into mice 1h before irinotecan administration (75 mg/kg, i.p.; 4 days), and then for additional 6 days. Seven days after the first dose of irinotecan, diarrhea was assessed and the intestine was removed for histological evaluation, assessment of intestinal over-contractility, measurement of myeloperoxidase (MPO), glutathione (GSH), malondialdehyde (MDA), proinflammatory cytokines and chemokine (IL-1, IL-6, and KC levels - a murine homolog of human IL-8 chemokine), analysis of cyclooxygenase 2 (COX-2), nuclear factor kappa B (NF-κB) and nitric oxide synthase (iNOS) expression. At the two highest doses, McLTP1 administration decreased mortality and diarrhea. McLTP1 (8 mg/kg, i.v.) significantly prevented irinotecan-induced intestinal damage and led to a reduction in over-contractility of the intestinal muscle (p < 0.05). Moreover, McLTP1 decreased the MPO, IL-1β, IL-6, and KC levels significantly by 74.7%, 42%, 92.9%, and 95.9%, respectively. Also, the expression of COX-2, NF-κB, and iNOS was reduced. Pretreatment with McLTP1 (8mg/kg; i.v.) significantly reduced the MDA level and increased the duodenum homogenates' GSH level. Our study provides a potential new therapeutic for preventing irinotecan-induced mucositis, improved clinical parameters, reduced inflammation, and oxidative damage.

Background: Calotropis procera is a laticiferous plant (Apocynaceae) found in tropical regions all over the world. The ultrastructural characteristics of laticifers, their restricted distribution among different taxonomic groups and in some species in each clade, as peptidases from latex, make them very attractive for biological analysis. Objective: To investigate the effects of LP-PII-IAA (laticifer protein (LP) sub-fraction II (PII) of C. procera presenting an iodoacetamide-inhibited cysteine proteinase activity) on irinotecan-induced intestinal mucositis, a serious adverse effect of this medicine for the treatment of cancer. Results: LP-PII-IAA is composed of closely related isoforms (90%) of peptidases deprived from catalysis and an osmotin protein (5%). Animals receiving co-administration of LP-PII-IAA presented a significant decrease in mortality, absence of diarrhea, histological preservation and normalization of intestinal functions. Clinical homeostasis was accompanied by a reduction in MPO activity and declined levels of IL-1β, IL-6 and KC while IL-10 level increased in LP-PII-IAA-treated animals. COX-2 and NF-kB immunostaining was reduced and the level of oxidative markers (GSH, MDA) were normalized in animals that received LP-PII-IAA. Conclusion: We suggest that peptidases from the latex of Calotropis procera were instrumental for the suppression of the adverse clinical and physiological effects of irinotecan.