Seroepidemiology of visceral leishmaniasis among free-roaming dogs and children in Zahedan city, southeast of Iran, 2018-2020.

BACKGROUND: Mediterranean visceral leishmaniasis (VL) or kala-azar is an endemic zoonotic disease in Iran. Domestic dogs are the primary reservoir host and source of VL infection. The high-risk populations are children and immune-deficient adults. OBJECTIVE: Based on the lack of published reports about the VL in Sistan and Baluchestan province in the southeast of Iran, this study aimed to assess the seroprevalence of diseae in free-roaming dogs and children under 12 years old using indirect fluorescent antibody (IFA) test. METHODS: This cross-sectional study was performed between 2018 and 2020 in Zahedan city, Sistan, and Baluchestan province. Blood samples were taken from 400 children under 12 years old with a fever history accompanied by at least another specific clinical presentation. In the same period, blood samples were collected from 150 stray dogs. Demographic characteristics and clinical manifestations in both humans and dogs were recorded. The IFA test examined all blood samples for the detection of anti-Leishmania infantum antibodies. RESULTS: Overall, the IFA test results were positive in 8 dogs (5.33%). Only two seropositive dogs (25%) showed obvious clinical symptoms. There was a significant correlation between the positive cases, clinical signs (P = 0.046), and age (P = 0.037) in infected dogs. None of the collected sera from 400 febrile children were positive. CONCLUSION: According to the present finding, it seems that VL is not endemic in Zahedan city, Sistan, and Baluchestan province, but the domestic cycle of L. infantum has been established in this area. Further investigations would be needed to estimate the status of VL infection in wild canines as a secondary potential reservoir host. Furthermore, periodic monitoring of disease must not be neglected.

Dirofilariosis caused by Dirofilaria immitis in the south of Kerman province, Iran.

BACKGROUND: Dirofilariosis due to Dirofilaria immitis is endemic in various areas of Iran. Domestic dogs are the main reservoirs and represent a major potential infection source for the vector and humans. OBJECTIVE: This study aimed to explore the prevalence of dirofilariosis due to D.immitis and its public health importance in domestic dogs in the Jiroft district, south of Kerman province, Iran, by serological and parasitological methods. METHODS: This descriptive study was carried out as a cross-sectional investigation. A questionnaire was completed for 100 domestic dogs from May 2017 to February 2018 and recorded their age, sex, and clinical features. Also, we used enzyme-linked immunosorbent assay (ELISA) to identify antigens of heartworms in the bloodstream, with 98% sensitivity and 100% specificity, and parasitological techniques (Knott's test) to detect microfilariae in canine blood in Jiroft district, south of Kerman province, Iran. RESULTS: Overall, 10 (10%) and 4 (4%) domestic dogs were infected as confirmed by ELISA and modified Knott's tests, respectively. The rate of occult infections in the ELISA test than Knott's test was 60%. No significant difference was found between dirofilariosis and gender. In contrast, there was a significant difference between dirofilariosis infection and age (P < 0.05). CONCLUSION: The present findings could help understand the epidemiological aspects of D. immitis for future control programs and take appropriate preventive and therapeutic strategies against the disease.

The impact of diabetes on cutaneous leishmaniasis: a case-control field assessment.

This study aimed to assess the associated-risk determinants for cutaneous leishmaniasis (CL) in patients with diabetes mellitus (DM) compared to patients without DM. This case-control study was performed between 2017 and 2019 in southeastern Iran. Overall, 206 participants were selected from patients with DM without CL (11.2%), patients with CL without DM (6.2%), and DM patients concomitance with CL (27.6%) as case groups and healthy individuals as a control group 64 (76%). These cases were compared for parasitological, immunological, biochemical, and hematological parameters. The findings demonstrated that parasitological factors regarding the number, duration, and size of the lesion in CL patients showed a significant difference among patients with and without DM (p < 0.05). Data analysis showed that six major risk factors, including female (odds ratio (OR) = 3.47, confidence interval (CI) = 1.84-6.53, p < 0.001), total protein in CL group (OR = 4.9, CI = 2.3-10.44, p < 0.001), alanine aminotransferase (ALT) concentration in CL group (OR = 0.87, CI = 0.81-0.93, p < 0.001) and DM co-infected with CL group (OR = 0.8, CI = 0.72-0.88, p < 0.001) than healthy group, aspartate aminotransferase (AST) concentration in DM group (OR = 0.86, CI = 0.76-0.98, p = 0.02), transforming growth factor beta)TGF-ß( level in the CL group (OR = 1.03, CI = 1.003-1.05, p = 0.02), and presence of diabetes disease (OR = 2.07, CI = 1.16-3.7, p < 0.05), were significantly linked with the induction of CL lesion. The findings demonstrated a significant relationship between DM and CL in distinct risk determinants. Also, the study revealed that DM enhanced the severity of active CL.

A single-group trial of end-stage patients with anthroponotic cutaneous leishmaniasis: Levamisole in combination with Glucantime in field and laboratory models.

Currently, there is no satisfactory treatment modality available for cutaneous leishmaniasis (CL). The major objective of the present study was to explore the effect of immunomodulator-levamisole in combination with Glucantime in end-stage unresponsive patients with anthroponotic CL (ACL). Twenty end-stage unresponsive patients with ACL were identified for participation in this single-group trial study. Simultaneously, each patient was received a combination of levamisole pills along with Glucantime during the remedy course. Several in vitro complementary experiments were performed to evaluate the mode of action of levamisole and Glucantime alone and in combination using a macrophage model, in vitro MTT assay, flow cytometry and quantitative real time PCR (qPCR). Overall, 75% of the patients showed complete clinical cure, 10% partially improved and the remaining (15%) had underlying chronic diseases demonstrated no response to the treatment regimen. In in vitro studies, there was no cytotoxic effect associated with these drugs in the range of our experiments. The findings by the flow cytometric analysis represented that the highest apoptotic values corresponded to the drugs combination (32.23%) at 200 µg/ml concentration. Finally, the gene expression level of IL-12 p40, iNOS and TNF-α promoted while the level of IL-10 and TGF-ß genes reduced as anticipated. The findings clearly indicated that the combination of levamisole and Glucantime should be considered in end-stage unresponsive patients with ACL who have not responded to basic treatments. The immunomodulatory role of levamisole in mounting immune system as documented by the in vitro experiments and further substantiated by this single-group trail study was highlighted.

A Novel Niosomal Combination of Selenium Coupled with Glucantime against Leishmania tropica.

There is no effective treatment modality available against different forms of leishmaniasis. Therefore, the aim of this study was to improve the penetration and efficacy of selenium and glucantime coupled with niosomes and compared them with their simple forms alone on in vitro susceptibility assays. In this study, the niosomal formulations of selenium and in combination with glucantime were prepared. The size and morphology of the niosomal formulations were characterized and the effectivity of the new formulation was also evaluated using in vitro MTT assay, intra-macrophage model, and gene expression profile. From the results obtained, no cytotoxicity effect was observed for niosomal and simple forms of drugs, as alone or in combination. Niosomal formulations of the drugs significantly showed more inhibitory effects (P ≤ 0.001) than the simple drugs when the selectivity index was considered. The gene expression levels of Interleukin (IL-10) significantly decreased, while the level of IL-12 and metacaspase significantly increased (P ≤ 0.001). The results of the present study showed that selenium plus glucantime niosome possess a potent anti-leishmanial effect and enhanced their lethal activity as evidenced by the in vitro experiments.

Feline visceral leishmaniasis in Kerman, southeast of Iran: Serological and molecular study.

BACKGROUND & OBJECTIVES: Visceral leishmaniasis (VL) is a fatal zoonotic disease in tropical and sub-tropical countries including Iran. Dogs constitute the main domestic reservoir for VL (kala-azar) in Iran but incidence of the disease in cats from Fars and East Azerbaijan provinces has led to propose them as secondary reservoirs, and possible expansion of the feline role in the transmission of disease. The aim of this study was to evaluate the prevalence of Leishmania infantum infection in stray cats in Kerman City by ELISA and PCR methods. METHODS: In this cross-sectional descriptive study, 60 stray cats were randomly live trapped from different parts of Kerman City during a six month period between March and September 2014. About 3 ml blood samples were drawn from jugular vein of captured cats and a detailed questionnaire about demographic characteristics and clini- cal status of each cat was recorded by attending veterinarian. The complete blood counts and biochemistry analysis were performed for all cats. Finally collected sera samples were tested by an indirect enzyme-linked immunosorbent assay (ELISA) kit and PCR amplification method. RESULTS: Prevalence of Leishmania infantum infection was 6.7 and 16.7% by ELISA and PCR assays, respectively. Infection rate was significantly higher in leukopenic cats, which were older than 3 yr. INTERPRETATION & CONCLUSION: The results of the study indicate that stray cats are at risk of L. infantum infection in Kerman City. Further, studies are required to elucidate the role of cats as potential reservoir host in the epidemiol- ogy of VL in endemic regions.

Effect of Administration of Lidocaine at Body Temperature on Anesthesia Success in Rodent Model: A Behavioral and Electrophysiology Study.

Introduction: Success in anesthesia administration relieves the perception of pain during surgery. Lidocaine is the most commonly used local anesthetic agent in clinical medicine. Moreover, anesthetic agents' temperature changes can influence cell membrane permeability. Here, the effectiveness of different temperatures of Lidocaine (Lid.) on anesthesia success rate has been investigated in rats. Methods: Wistar male rats were pretreated by fast injection of lidocaine or saline into the hind paw or intradermal cheek at Room Temperature (RT) and Body Temperature (BT) (22°C and 37°C, respectively). Then, rat behaviors were evaluated by formalin-induced hind paw pain and orofacial pain tests, respectively. Moreover, using a single-unit recording technique, the spontaneous activity of the marginal nerve was recorded at room temperature in the RT-Lid. and BT-Lid. groups. Results: Data analysis revealed that lidocaine had significant antinociceptive effects in both the BT-Lid. and RT-Lid. groups compared to the control groups (P<0.05). Also, the number of spikes in the BT-Lid. and RT-Lid. groups were significantly lower than their baselines (P<0.05). However, lidocaine at body temperature decreased the total time spent licking the hind paw, the number of lip rubbings, and the number of spikes firing by about 10%-15% compared to room temperature. Conclusion: In both behavioral and neural levels of the study, our results showed that an increase in the temperature of lidocaine toward body temperature could increase anesthesia success rate compared to administration of lidocaine at room temperature. These findings can be considered in the treatment of patients. Highlights: Lidocaine at body temperature acted better than room temperature on pain control in the formalin-induced hind paw test.Lidocaine at body temperature acted better than room temperature on pain control in the orofacial formalin test.Lidocaine with different temperatures decreased the firing rate of the marginal nerve. Plain Language Summary: Pain is defined as an unpleasant experience caused by tissue damage or fear of injury. During anesthetic injection in dentistry, pain has long been one of the problems of dentists. Studies have shown that one out of every three people is worried about going to dentistry, and one of four dental patients is afraid of injections. The fear of a patient in one of twenty patients is so much that interferes with dental treatment which consequently leads to stress when you visit the dentist, results in less oral hygiene and reduces the number of referrals. Lidocaine is the most commonly used local anesthetic agent in clinical medicine. Here, the effectiveness of different temperatures of lidocaine on anesthesia success rate in rats has been investigated in rats. The present study showed that warming the lidocaine cartridges to 37°C increased anesthesia success compared to anesthesia-induced at room temperature in both behavioral and neural levels of the study. Accordingly, a warmed anesthetic cartridge could be used to control pain by increasing the success rate during dental injection and designed a new animal model study for further investigation in comparing other anesthesia drugs.

Antiproliferative properties of Turmerone on Leishmania major: Modes of action confirmed by antioxidative and immunomodulatory roles.

Treatment of leishmaniasis by conventional synthetic compounds has faced a serious challenge worldwide. This study was performed to evaluate the effect and modes of action of aromatic Turmerone on the Leishmania major intra-macrophage amastigotes, the causative agent of zoonotic cutaneous leishmaniasis in the Old World. In the findings, the mean numbers of L. major amastigotes in macrophages were significantly decreased in exposure to Turmerone plus meglumine antimoniate (Glucantime®; MA) than MA alone, especially at 50 µg/mL. In addition, Turmerone demonstrated no cytotoxicity as the selectivity index (SI) was 21.1; while it induced significant apoptosis in a dose-dependent manner on L. major promastigotes. In silico molecular docking analyses indicated an affinity of Turmerone to IL-12, with the MolDock score of - 96.8 kcal/mol; which may explain the increased levels of Th1 cytokines and decreased level of IL-10. The main mechanism of action is more likely associated with stimulating a powerful antioxidant and promoting the immunomodulatory roles in the killing of the target organism.

The Fraction of the Snake Venom, Its Leishmanicidal Effect, and the Stimulation of an Anti-Leishmania Response in Infected Macrophages.

BACKGROUND AND AIMS: Due to the lack of an effective vaccine and complexity of the control measures against vectors and reservoir hosts, the control of leishmaniasis depends primarily on chemotherapy. This study was aimed to assess the snake venom, Naja naja oxiana fraction 11(NNOVF11) on Leishmania infantum and its broad mode of action. METHODS: A wide range of in vitro advanced assays including high-performance liquid chromatography (HPLC), MTT (3-[4, 5-Dimethylthiazol-2-yl]-2, 5diphenyltetrazolium bromide; Thiazolyl blue), macrophage assays, quantitative real-time polymerase chain reaction (qPCR), flow cytometry and enzyme- linked immunosorbent assay (ELISA) on L. infantum promastigote and amastigote stages were used. IC50 values of L. infantum stages, CC50 value, and apoptosis were also analyzed. RESULTS: The NNOV-F11 demonstrated strong antileishmanial activity against L. infantum stages in a dose-dependent manner compared to the untreated control group. Interleukin (IL)-12, TNF-α, and iNOS genes expression as the indicators of T helper(h)1 response significantly increased; in contrast, the expression level of IL-10, as the representative of Th2 response significantly decreased (p < 0.001). Reactive oxygen species (ROS) detection showed a significant increase (p < 0.001) after treatment with different concentrations of NNOV-F11, unlike arginase (ARG) activity, which displayed a significant reduction (p < 0.001). CONCLUSION: NNOV-F11 possessed a potent inhibitory effect on L. infantum stages with the multifunctional and broad mode of actions, which promoted the immunomodulatory role, induced ROS production, stimulated apoptotic-like mechanisms, and inhibited L-ARG activity, which collectively led to the parasite death. Further studies are crucial to assess the effect of the NNOV-F11 on animal models or clinical settings.

The Effect of Naja naja oxiana Snake Venom Against Leishmania tropica Confirmed by Advanced Assays.

PURPOSE: The aim of this study was to explore the activity of Naja naja oxiana venom on Leishmania tropica and its modes of action. METHODS: Different fractions of Naja naja oxiana venom (NNOV) were prepared and characterized by high-performance liquid chromatography. The superior component, fraction k (FK) was selected. The activity of the fraction was assessed using advanced assays. RESULTS: Interleukin (IL)-12, TNF-α and iNOS gene expression as the indicators of Th1 significantly increased. In contrast, the level of IL-10, as the marker of T helper 2 substantially decreased (p < 0.001). Reactive oxygen species (ROS) detection showed a significant increase (p < 0.001) after treatment with different concentrations of NNOV-FK, unlike arginase (L-ARG) activity which showed a significant reduction (p < 0.001). The NNOV-FK showed significant lethal activity on the L. tropica stages. CONCLUSION: The findings demonstrated that NNOV-FK represented a strong leishmanicidal activity on L. tropica stages. The major modes of NNOV-FK action are multidimensional, which perceives the induction of a synergistic response and upregulation of the immune-modulatory role towards Th1 response against L. tropica stages as well as apoptotic and anti-metabolic action as a model drug to generate ROS, block the polyamine synthesis and lead to parasite death.

The Knowledge and Attitude of Diabetic Patients Regarding Oral and Dental Disorders in Kerman Diabetes Clinics.

STATEMENT OF THE PROBLEM: Diabetes mellitus is one of the most common endocrine disorders. This disease has devastating effects on many organs and tissues of the body including oral and dental tissues. PURPOSE: The aim of this study was to evaluate the knowledge and attitude of diabetic patients about dental and oral diseases. MATERIALS AND METHOD: In this cross-sectional study, 433 diabetic patients who referred to Kerman Diabetes Clinics were included. Data were collected using a questionnaire consisted of three parts of demographic characteristics, knowledge of oral and systemic complications of diabetes mellitus, and patients' attitude regarding their oral health. Data were analyzed using SPSS version 21 and employing t-test and multiple linear regression analysis. Statistically significant values were considered at p≤ 0.05. RESULTS: The mean scores for the knowledge of systemic and oral complications were 0.80±0.21 and 0.39±0.23, respectively. The mean total knowledge of diabetic patients was 0.53±0.18, and the mean score for the patients' attitude was 0.63±0.11. It was revealed that people with a family history of diabetes did not have significantly greater overall knowledge (p= 0.082). Also, people with longer disease duration (p= 0.004) and female patients (p= 0.05) had significantly a better knowledge and attitude in terms of oral health. CONCLUSION: The knowledge and attitude of patients regarding their oral and dental health and diseases were at moderate level, which should be promoted by constant planning and education according to the current needs of society.

Linguatula serrata in cattle in southeastern Iran: Epidemiological, histopathological and phylogenetic profile and its zoonotic importance.

This study aimed to determine the prevalence, histopathological observations, and phylogenetic analysis of L. serrata in cattle and its potential zoonotic and public health implications in southeastern Iran.The cross-sectional study was performed in cattle in southeastern Iran. Lymph nodes were collected from each cattle and examined by parasitological and histopathological techniques. A binary logistic regression and chi-square tests were implemented to analyze the data. Genomic DNA was randomly extracted from the nymphal stages of Linguatula isolates. Further characterization and phylogenetic relationships were done using two primers for amplification of partial DNA fragments of 18 s rRNA and cytochrome C oxidase subunit 1 (cox1), respectively.The results showed that 64 cattle of the total 404 were infected with L. serrata. There was no significant difference between linguatulosis infection and gender, while age was significantly different (P < 0.05). The cyst-like spaces containing the longitudinal and transverse sections of the L. serrata nymphs were surrounded by granulomatous reactions. The higher nucleotide variation in the cox1 region was supported by estimating the evolutionary divergence between L. serrata isolates and other Linguatula records of ruminants in Iran. The phylogenetic tree confirmed the close evolutionary relationships among all reported records of L. serrata in Iran.The high prevalence of linguatulosis caused by L. serrata declares the existence of a potential risk of FBPs for humans in southeastern Iran. This condition can advance more serious public health problems and requires a comprehensive control program and treatment strategies to prevent the disease.

The potential role of nicotinamide on Leishmania tropica: An assessment of inhibitory effect, cytokines gene expression and arginase profiling.

Leishmaniasis represents a major health concern worldwide which has no effective treatment modality. Nicotinamide (NAm) has been used for a wide range of applications from anticancer to antimicrobial usage. This study aimed to assess the effect of NAm combination on Leishmania tropica Inhibition, as well as on cytokines gene expression and arginase (ARG) activity in L. tropica-infected macrophages in an in vitro model. The leishmanicidal effects of NAm and Glucantime (meglumine antimoniate, MA) alone and in combination (NAm/MA) were evaluated using a colorimetric assay and macrophage model. Additionally, immunomodulatory effects and enzymatic activity were assessed by analyzing Th1 and Th2 cytokines gene expression and ARG level, respectively, in infected macrophages treated with NAm and MA, alone and in combination. Findings indicated that the NAm/MA combination demonstrated greater inhibitory effects on L. tropica promastigotes and amastigotes compared with each drug individually. Docking results proved the affinity of NAm to IFN-γ, which can affirm the increased levels of IFN-γ, IL-12p40 and TNF-α as well as reductions in IL-10 secretion with a dose-response effect, especially in the combination group. The NAm/MA combination also showed a significant reduction in the level of ARG activity at all concentrations used compared to each drug individually. These findings indicate higher effectiveness of NAm plus MA in reducing parasite growth, promoting immune response and inhibiting ARG level. This combination should be considered as a potential therapeutic regimen for treatment of volunteer patients with anthroponotic cutaneous leishmaniasis (ACL) in future control programs.

Leishmanicidal effects of amphotericin B in combination with selenium loaded on niosome against Leishmania tropica.

The strategy for improving the treatment of leishmaniasis by the World Health Organization, is the development of new drugs and combination therapy. The aim of this survey was to investigate the effect of amphotericin B (AmB) in combination with selenium, in a simple or niosomal form, on Leishmania tropica (L. tropica) by in vitro advanced assays. In this study, a niosomal formulation of AmB with selenium was prepared and characterized based on size and morphology. Using MTT assay, macrophage model, flow cytometry, and qPCR, the cytotoxicity and efficiency of the niosomal formulation and simple form of combination were evaluated. No toxicity was reported for both the niosomal and simple form of the combination. The niosomal formulation significantly showed higher inhibitory effect on the promastigote and amastigote forms of L. tropica than simple combination form. Interleukin (IL)-10 significantly decreased while the level of IL-12 and metacasoase as Th-1 activator significantly increased (P < 0.001). The findings of this study indicated that niosomes are the stable carriers for this combination, easy to produce and provide promising results as an effective formulation in the inhibition of extracellular and intracellular forms of L. tropica in compared with simple combination form.

Antileishmanial activity and immune modulatory effects of benzoxonium chloride and its entrapped forms in niosome on Leishmania tropica.

Benzoxonium chloride is an anti-infective agent that is used as anti-septic drugs for disinfection of the mucus membrane, skin surface and anti-bacterial, and it is also found to be effective against cutaneous leishmaniasis. The present study aims to evaluate the leishmanicidal activity of benzoxonium chloride and niosomal forms against Leishmania tropica stages. Benzoxonium chloride niosomes were prepared by the thin film hydration method and evaluated for morphology, particle size and release study and encapsulation efficiency. This study measured the cytotoxicity, leishmanicidal activity against promastigote and intra macrophage amastigote, apoptosis, and mRNA transcripts by quantitative real time PCR (qPCR) of free solution and niosomal-encapsulated benzoxonium chloride. Span/Tween 60 niosomal formulation of benzoxonium chloride showed superior physical stability and high encapsulation efficiency (96%) than the other forms. Release from the formulations showed that the Span/Tween 60 containing drug had a milder gradient so that 10% of the drug was not released after 4 h. The benzoxonium chloride and niosomal forms inhibited the in vitro growth of promastigote and amastigote forms of L. tropica after 48 h of incubation and represented IC50 values of 90.7 ± 2.7 and 25.4 ± 0.6 µg/ mL, respectively. The rate of apoptosis in niosomal formulations was approximately equal to the positive control (meglumine antimoniate) at the same concentration. Also, an increase in the concentration of this drug reduced the expression of IL-10, but increased the expression of IL-12. The niosomal formulations provided improved anti-leishmanial activities of benzoxonium chloride and played an immunomodulatory role as the mode of action in the treatment of anthroponotic CL.

Host's immune response in unresponsive and responsive patients with anthroponotic cutaneous leishmaniasis treated by meglumine antimoniate: A case-control study of Th1 and Th2 pathways.

The objective of the present study was to compare the host's immune responses between unresponsive and responsive patients with anthroponotic cutaneous leishmaniasis (ACL) treated by meglumine antimoniate. A case-control study was carried out in an endemic focus in Iran. Blood samples were taken from patients and peripheral blood mononuclear cells (PBMCs) were isolated. Two wells were considered for each isolate of unresponsive and responsive patients; one was exposed to L. tropica (Lt-stimulated cells) and the other remained non-exposed (non-stimulated cells). After 24 h of incubation, whole RNA was extracted from each sample. Real-time quantitative PCR was carried out to confirm the differences in expression levels of IL-12 P40, IFN-γ, IL-1ß, IL-4 and IL-10 among isolates. Data were analyzed and P < 0.05 was considered to be statistically significant. In our study, Lt-stimulated cells and non-stimulated cells in unresponsive groups demonstrated significantly lower expression levels of IL-1ß, IL-12 P40 and IFN-γ genes and higher expression levels of IL-4 and IL-10 genes, compared to Lt-stimulated cells and non-stimulated cells in responsive groups. There was a negative correlation between IL-12 P40 with IL-10 and IL-1ß with IL-10 in ACL Lt-stimulated cells in unresponsive group, while a positive correlation between IL-12 P40 with IL-1ß and IL-12 P40 with IFN-γ in ACL Lt-stimulated cells in responsive group. Probably, different immune responses caused by various factors play a major role in the pathogenesis and development of unresponsiveness in ACL patients. The profile and timing of cytokine production correlated well with the treatment outcome of Leishmania infection.

Niosomal formulation of amphotericin B alone and in combination with glucantime: In vitro and in vivo leishmanicidal effects.

This study aimed to evaluate the efficacy of glucantime and amphotericin B (AmB) encapsulated in niosome against cutaneous leishmaniasis (CL) using in vitro and in vivo models. The niosomal formulations of the drugs alone and in combination were prepared and characterized. Subsequent to the examination of their cytotoxicity, their efficacy was evaluated using an in vitro MTT assay, macrophage model, flow cytometry, and gene expression profiling. For evaluation of therapeutic effect of niosomal combination on the lesion induced by Leishmania major in inbred BALB/c mice, the size of lesions and number of parasites in spleen was assessed. The niosomal formulations demonstrated significantly greater inhibitory effects compared with the non-niosomal forms when the IC50 was considered. The niosomal combination showed an increase in the apoptotic values and gene expression levels of IL-12 and metacaspase and a decrease in the levels of IL-10 with a dose-response effect. The niosomal combination was also effective in reducing the lesion size and splenic parasite burden in mice. Our findings indicated that there is a synergistic effect between AmB and glucantime in niosomal form in the inhibition of intracellular and extracellular forms of L. tropica. Additionally, the in vivo results on L. major suggest that topical niosomal formulation could be useful in the treatment of CL.

Toxico-pathological effects of meglumine antimoniate on human umbilical vein endothelial cells.

Leishmaniasis is one of the most important parasitic diseases after malaria. The standard treatment of leishmaniasis includes pentavalent antimonials (SbV); however, these drugs are associated with serious adverse effects. There have been very few studies pertaining to their side effects and mechanism of action in the fetus. This investigation examines the effects of meglumine antimoniate (MA) on the survival rate, angiogenesis and cellular apoptosis in the human umbilical vein endothelial cells (HUVECs). HUVECs were treated with varying doses of MA (100-800 µg/ml) for 24, 48 and 72 h and the survival rate was studied by colorimetric assay, flow cytometry, immunocytochemistry, migration (scratch) assay and tube formation assay. The results of quantitative real-time PCR (qPCR) studies indicated that the most important genes involved in presenting angiogenesis included VEGF and its receptors (Kdr and Flt-1), NP1 and Hif-1α genes including the anti-apoptotic gene of Bcl2, were significantly reduced compared to the control group (p < 0.05). In contrast, the most leading genes involved in the phenomenon of apoptosis were P53, Bax, Bak, Apaf-1 and caspases 3, 8 and 9, which were significantly up regulated compared to the control group (p < 0.05).

Anti-Leishmanial and Immunomodulatory Effects of Epigallocatechin 3-O-Gallate on Leishmania tropica: Apoptosis and Gene Expression Profiling.

BACKGROUND: Pentavalent antimonials such as meglumine antimoniate (MA, Glucantime), are the first-line treatment against leishmaniasis, but at present, they have basically lost their efficacy. This study was aimed to explore epigallocatechin 3-O-gallate (EGCG), alone or in combination with MA against Leishmania tropica stages. METHODS: All experiments were carried out in triplicate using colorimetric assay, macrophage model, flow cytometry and quantitative real-time PCR. This experimental study was carried out in 2017 in Leishmaniasis Research Center, Kerman University of Medical Sciences, Kerman, Iran. RESULTS: Promastigotes and amastigotes were more susceptible to EGCG than MA alone, but the effect was more profound when used in combination. EGCG exhibited high antioxidant level with a remarkable potential to induce apoptosis. Furthermore, the results showed that the level of gene expression pertaining to Th-1 was significantly up-regulated (P<0.001). CONCLUSION: EGCG demonstrated a potent anti-leishmanial effect alone and more enhanced lethal activity in combination. The principal mode of action entails the stimulation of a synergistic response and up-regulation of the immunomodulatory role towards Th-1 response against L. tropica.

Clinical Features of Anthroponotic Cutaneous Leishmaniasis in a Major Focus, Southeastern Iran, 1994-2014.

BACKGROUND: Cutaneous leishmaniasis (CL) is associated with a broad and complex clinical spectrum of diseases. The objectives of this study were to assess the clinical features and identification of the causative agents of CL in a well-known focus of anthroponotic CL (ACL) caused by Leishmania tropica, southeast Iran. METHODS: This study was performed randomly as a descriptive cross-sectional survey to evaluate 2000 CL patients by active and passive case-detection approaches in Kerman Province from 1994 to 2014. The ACL patients were confirmed by direct smear and 600 cases by one or a combination of intrinsic methods. RESULTS: Children aged <10 yr old were the most infected patients (P<0.001). The majority of the CL lesions were located in hands (46.3%), face (34.1%), legs (14.3%), and other parts of the body (5.3%). The mean number of lesions was 1.5 and most of the patients had single lesion (65%).Typical clinical lesions included papule (36.8%), followed by ulcerated nodule (20.7%), plaque (18.4%), and ulcerated plaque (18.5%). While among atypical clinical features, leishmaniasis recidivans (LR) (4.7%) and leishmanid (0.3%) were the dominant forms, followed by diffuse, disseminated, sporotrichoid, and erysipeloid types, 0.1% each, and then lymphedematous, lymphadenic, hyperkeratotic, paronychial, and mutilating types, 0.05% each. Based on various intrinsic methods the parasites isolated from the lesions were characterized as L. tropica. CONCLUSION: ACL due to L. tropica presents numerous cases of localized form and diverse uncommon clinical presentations, which mimic other disease conditions. Therefore, physicians should be aware of such manifestations for selecting appropriate treatment modality.