Immunophenotyping of Peripheral Blood, Lymph Node, and Bone Marrow T Lymphocytes During Canine Leishmaniosis and the Impact of Antileishmanial Chemotherapy.

Dogs are a major reservoir of Leishmania infantum, etiological agent of canine leishmaniosis (CanL) a zoonotic visceral disease of worldwide concern. Therapeutic protocols based on antileishmanial drugs are commonly used to treat sick dogs and improve their clinical condition. To better understand the impact of Leishmania infection and antileishmanial drugs on the dog's immune response, this study investigates the profile of CD4+ and CD8+ T cell subsets in peripheral blood, lymph node, and bone marrow of sick dogs and after two different CanL treatments. Two CanL groups of six dogs each were treated with either miltefosine or meglumine antimoniate combined with allopurinol. Another group of 10 clinically healthy dogs was used as control. Upon diagnosis and during the following 3 months of treatment, peripheral blood, popliteal lymph node, and bone marrow mononuclear cells were collected, labeled for surface markers CD45, CD3, CD4, CD8, CD25, and intracellular nuclear factor FoxP3, and T lymphocyte subpopulations were immunophenotyped by flow cytometry. CanL dogs presented an overall increased frequency of CD8+ and CD4+CD8+ double-positive T cells in all tissues and a decreased frequency of CD4+ T cells in the blood. Furthermore, there was a higher frequency of CD8+ T cells expressing CD25+FoxP3+ in the blood and bone marrow. During treatment, these subsets recovered to levels similar to those of healthy dogs. Nevertheless, antileishmanial therapy caused an increase of CD4+CD25+FoxP3+ T cells in all tissues, associated with the decrease of CD8+CD25-FoxP3- T cell percentages. These findings may support previous studies that indicate that L. infantum manipulates the dog's immune system to avoid the development of a protective response, ensuring the parasite's survival and the conditions that allow the completion of Leishmania life cycle. Both treatments used appear to have an effect on the dog's immune response, proving to be effective in promoting the normalization of T cell subsets.

Meglumine Antimoniate and Miltefosine Combined With Allopurinol Sustain Pro-inflammatory Immune Environments During Canine Leishmaniosis Treatment.

Canine leishmaniosis (CanL) caused by Leishmania infantum is a zoonotic disease of global concern. Antileishmanial drug therapies commonly used to treat sick dogs improve their clinical condition, although when discontinued relapses can occur. Thus, the current study aims to evaluate the effect of CanL treatments in peripheral blood, lymph node, and bone marrow cytokine profile associated with clinical recovery. Two groups of six dogs diagnosed with CanL were treated with miltefosine combined with allopurinol and meglumine antimoniate combined with allopurinol (MT+A and MG+A), respectively. At diagnosis and after treatment, during a 3-month follow-up, clinical signs, hematological and biochemical parameters, urinalysis results and antileishmanial antibody titers were registered. Furthermore, peripheral blood, popliteal lymph node, and bone marrow samples were collected to assess the gene expression of IL-2, IL-4, IL-5, IL-10, IL-12, TNF-α, TGF-ß, and IFN-γ by qPCR. In parallel, were also evaluated samples obtained from five healthy dogs. Both treatment protocols promoted the remission of clinical signs as well as normalization of hematological and biochemical parameters and urinalysis values. Antileishmanial antibodies returned to non-significant titers in all dogs. Sick dogs showed a generalized upregulation of IFN-γ and downregulation of IL-2, IL-4, and TGF-ß, while gene expression of IL-12, TNF-α, IL-5, and IL-10 varied between groups and according to evaluated tissue. A trend to the normalization of cytokine gene expression was induced by both miltefosine and meglumine antimoniate combined therapies. However, IFN-γ gene expression was still up-regulated in the three evaluated tissues. Furthermore, the effect of treatment in the gene expression of cytokines that were not significantly changed by infection, indicates that miltefosine and meglumine antimoniate combined therapy directly affects cytokine generation. Both combined therapies are effective in CanL treatment, leading to sustained pro-inflammatory immune environments that can compromise parasite survival and favor dogs' clinical cure. In the current study, anti-inflammatory and regulatory cytokines do not seem to play a prominent role in CanL or during clinical recovery.

Análise da Implantação do Protocolo para Incorporaçãode Pacientes de Grupos Prioritários na Atenção Básica / Analysis of Implementation of the Protocol for Incorporation of Patients from the Priority Groups Primary Health Care

Relata a experiência do município de Santana de Parnaíba (SP) para reverter à porta de entrada no sistema de saúde com ampliação do acesso às unidades básicas de saúde, em especial para grupos populacionais identificados como prioritários. Descreve os passos dados para este fim, assim como os instrumentos criados para a implantação do protocolo e seu monitoramento. A análise da participação das consultas de urgência/emergência, no período estudado frente ao total de consultas, mostra uma queda expressiva, passando de 76,96% no ano de 2000 para 53,18% em 2007. No mesmo período, as consultas em atenção básica sofreram incremento de aproximadamente 100%. Analisa as ações desenvolvidas pelas unidades de saúde ao receberem o paciente encaminhado pelas unidades socorristas, verificando ser a implantação do acolhimento uma realidade que reorganiza o sistema a partir das necessidadesde saúde e não apenas por critérios burocráticos e administrativos.

This article aims to report the experience of the municipality ofSantana de Parnaíba - SP, to revert the port of entry to the health system withexpansion of access to primary health, especially for population groups identifiedas priorities. At this work are described the steps taken to this end, and theinstruments created for the implementation of the protocol and its monitoring.The analysis of the participation of emergency / rescue consultations in front ofthe hole consultations from the full study period, shows a significant decline, from76.96% in 2000 to 53.18% in 2007. In the same period, the consultations inprimary health care suffered increase of approximately 100%. It analysis theactions undertaken by units of health when received patients sent emergencycenters, observing that the implantation of a service all patients in primaryhealth centers is a reality on the reorganization of the system, from the needs ofhealth and not just by bureaucratic and administrative criteria.