Improved Treatment Outcome Following the Use of a Wound Dressings in Cutaneous Leishmaniasis Lesions.

Leishmaniasis, caused by Leishmania parasites, is a neglected tropical disease and Cutaneous Leishmaniasis (CL) is the most common form. Despite the associated toxicity and adverse effects, Meglumine antimoniate (MA) remains the first-choice treatment for CL in Brazil, pressing the need for the development of better alternatives. Bacterial NanoCellulose (BNC), a biocompatible nanomaterial, has unique properties regarding wound healing. In a previous study, we showed that use of topical BNC + systemic MA significantly increased the cure rate of CL patients, compared to treatment with MA alone. Herein, we performed a study comparing the combination of a wound dressing (BNC or placebo) plus systemic MA versus systemic MA alone, in CL caused by Leishmania braziliensis. We show that patients treated with the combination treatment (BNC or placebo) + MA showed improved cure rates and decreased need for rescue treatment, although differences compared to controls (systemic MA alone) were not significant. However, the overall time-to-cure was significantly lower in groups treated with the combination treatment (BNC+ systemic MA or placebo + systemic MA) in comparison to controls (MA alone), indicating that the use of a wound dressing improves CL treatment outcome. Assessment of the immune response in peripheral blood showed an overall downmodulation in the inflammatory landscape and a significant decrease in the production of IL-1a (p < 0.05) in patients treated with topical BNC + systemic MA. Our results show that the application of wound dressings to CL lesions can improve chemotherapy outcome in CL caused by L. braziliensis.

Immunization with centrin-Deficient Leishmania braziliensis Does Not Protect against Homologous Challenge.

Immunization with various Leishmania species lacking centrin induces robust immunity against a homologous and heterologous virulent challenge, making centrin mutants a putative candidate for a leishmaniasis vaccine. Centrin is a calcium-binding cytoskeletal protein involved in centrosome duplication in higher eukaryotes and Leishmania spp. lacking centrin are unable to replicate in vivo and are non-pathogenic. We developed a centrin-deficient Leishmania braziliensis (LbCen-/-) cell line and confirmed its impaired survival following phagocytosis by macrophages. Upon experimental inoculation into BALB/c mice, LbCen-/- failed to induce lesions and parasites were rapidly eliminated. The immune response following inoculation with LbCen-/- was characterized by a mixed IFN-γ, IL-4, and IL-10 response and did not confer protection against L. braziliensis infection, distinct from L. major, L. donovani, and L mexicana centrin-deficient mutants. A prime-boost strategy also did not lead to a protective immune response against homologous challenge. On the contrary, immunization with centrin-deficient L. donovani (LdonCen-/-) cross-protected against L. braziliensis challenge, illustrating the ability of LdonCen-/- to induce the Th1-dominant protective immunity needed for leishmaniasis control. In conclusion, while centrin deficiency in L. braziliensis causes attenuation of virulence, and disrupts the ability to cause disease, it fails to stimulate a protective immune response.

A pilot and open trial to evaluate topical Bacterial Cellulose bio-curatives in the treatment of cutaneous leishmaniasis caused by L. braziliensis.

The treatment of cutaneous leishmaniasis (CL) in Brazil using pentavalent antimony (Sbv) is associated with a high failure rate and long time to heal. Moreover, standard Sbv treatment cures only 50-60% of the cases. In this pilot clinical trial, we evaluated the topical use of bacterial cellulose (BC) bio-curatives + Sbv in the treatment of CL caused by L. braziliensis, in Bahia, Brazil. A total of 20 patients were randomized in two groups assigned to receive either parenteral Sbv alone or parenteral Sbv plus topically applied BC bio-curatives. CL patients treated with Sbv + topical BC bio-curatives had a significantly higher cure rate at 60 days post initiation of treatment compared to CL patients treated with Sbv alone (P=0.01). At day 90 post initiation of treatment, cure rate was similar in the two groups as was overall healing time. Adverse effects or local reactions to topical BC application were not observed. This pilot trial shows that the potential use of a combined therapy consisting of topical BC bio-curatives and parenteral Sbv in favoring healing of CL lesions caused by L. braziliensis, at an early time point.

Targeted Deletion of Centrin in Leishmania braziliensis Using CRISPR-Cas9-Based Editing.

Leishmania braziliensis is the main causative agent of Tegumentary Leishmaniasis in the Americas. However, difficulties related to genome manipulation, experimental infection, and parasite growth have so far limited studies with this species. CRISPR-Cas9-based technology has made genome editing more accessible, and here we have successfully employed the LeishGEdit approach to attenuate L. braziliensis. We generated a transgenic cell line expressing Cas9 and T7 RNA polymerase, which was employed for the targeted deletion of centrin, a calcium-binding cytoskeletal protein involved in the centrosome duplication in eukaryotes. Centrin-deficient Leishmania exhibit growth arrest at the amastigote stage. Whole-genome sequencing of centrin-deficient L. braziliensis (LbCen-/- ) did not indicate the presence of off-target mutations. In vitro, the growth rates of LbCen-/- and wild-type promastigotes were similar, but axenic and intracellular LbCen-/- amastigotes showed a multinucleated phenotype with impaired survival following macrophage infection. Upon inoculation into BALB/c mice, LbCen-/- were detected at an early time point but failed to induce lesion formation, contrary to control animals, infected with wild-type L. braziliensis. A significantly lower parasite burden was also observed in mice inoculated with LbCen-/- , differently from control mice. Given that centrin-deficient Leishmania sp. have become candidates for vaccine development, we propose that LbCen-/- can be further explored for the purposes of immunoprophylaxis against American Tegumentary Leishmaniasis.

An exergame system based on force platforms and body key-point detection for balance training.

Postural instability affects a large number of people and can compromise even simple activities of the daily routine. Therapies for balance training can strongly benefit from auxiliary devices specially designed for this purpose. In this paper, we present a system for balance training that uses the metaphor of a game, what contributes to the motivation and engagement of the patients during a treatment. Such approach is usually named exergame, in which input devices for posturographic assessment and a visual output perform the interaction with the subject. The proposed system uses two force platforms, one positioned under the feet and the other under the hip of the subject. The force platforms employ regular load cells and a microcontroller-based signal acquisition module to capture and transmit the samples to a computer. Moreover, a computer vision module performs body key-point detection, based on real time segmentation of markers attached to the subject. For the validation of the system, we conducted experiments with 20 neurologically intact volunteers during two tests: comparison of the stabilometric parameters obtained from the system with those obtained from a commercial baropodometer and the practice of several exergames. Results show that the proposed system is completely functional and can be used as a versatile tool for balance training.

In a methotrexate-induced model of intestinal mucositis, olmesartan reduced inflammation and induced enteropathy characterized by severe diarrhea, weight loss, and reduced sucrose activity.

The aim of this study was to evaluate the effect of olmesartan (OLME), an angiotensin II receptor antagonist, on an intestinal mucositis model. Briefly, daily intraperitoneal (i.p.) injections of methotrexate (MTX) 7 mg/kg were administered to rats on 3 consecutive days. A subset of these rats was also pretreated with oral administration of OLME (0.5, 1.0, or 5.0 mg/kg) or vehicle as a control 30 min prior to MTX injection. Body weight, feces scoring, and death were recorded daily. On day 4, the rats were killed, and intestinal tissues were assayed for levels of interleukin (IL)-1ß and tumor necrosis factor (TNF)-α, myeloperoxidase and sucrose activity, and histopathological findings. A significant reduction in body weight was observed in the MTX+1.0 mg/kg OLME group (p<0.01). The feces scores for the MTX+0.5 mg/kg OLME and MTX+5.0 mg/kg OLME groups were also significantly higher (p<0.001). Sucrose activity was reduced in all groups treated with OLME (p<0.05). Treatment with MTX+OLM at all doses resulted in reduced inflammatory infiltration, ulcerations, vasodilation, and hemorrhagic areas (p<0.05), as well as reduced concentrations of myeloperoxidase (p<0.001). The IL-1ß and TNF-α levels were decreased in the MTX+OLME 5.0 mg/kg (p<0.01 and p<0.05, respectively) compared with the MTX-alone group. Overall, antiinflammatory activity was observed in rats with MTX-induced intestinal mucositis that were administered OLME. However, further studies are needed to elucidate the adverse effects of OLME.