Fluorescent membrane markers elucidate the association of Borrelia burgdorferi with tick cell lines

This study aimed to describe the association of Borrelia burgdorferi s.s. with ixodid tick cell lines by flow cytometry and fluorescence and confocal microscopy. Spirochetes were stained with a fluorescent membrane marker (PKH67 or PKH26), inoculated into 8 different tick cell lines and incubated at 30°C for 24 h. PKH efficiently stained B. burgdorferi without affecting bacterial viability or motility. Among the tick cell lines tested, the Rhipicephalus appendiculatus cell line RA243 achieved the highest percentage of association/internalization, with both high (90%) and low (10%) concentrations of BSK-H medium in tick cell culture medium. Treatment with cytochalasin D dramatically reduced the average percentage of cells with internalized spirochetes, which passed through a dramatic morphological change during their internalization by the host cell as observed in time-lapse photography. Almost all of the fluorescent bacteria were seen to be inside the tick cells. PKH labeling of borreliae proved to be a reliable and valuable tool to analyze the association of spirochetes with host cells by flow cytometry, confocal and fluorescence microscopy.

Effect of the entomopathogenic nematode Steinernema Glaseri (Rhabditida: Steinernematidae) isolate Santa Rosa on the biological parameters of engorged nymphs of Amblyomma Cajennense (Acari: Ixodidae) / Os efeitos do nematoide entomopatogênico steinernema glaseri (rhabditida: steinernematidae) isolado Santa Maria sobre os parâmetros Biológicos de ninfas ingurgitadas de amblyomma cajennense (acari: Ixodidae)

This work evaluated the in vitro effects of Steinernema glaseri on the biology of engorged Amblyomma cajennense nymphs. The ticks were exposed to infective juveniles (IJs) of S. glaseri at different concentrations. Each treatment was composed by 10 experimental units (EU = repetitions) in Petri dishes maintained at 27 ± 1º C and relative humidity (RH) > 80 ± 10% for 72 hours. Then the nymphs were transferred to Petri dishes free of nematodes for observation of the following parameters: pre-ecdysis period (PEP), ecdysis period (EP), molting period (MP), ecdysis percentage (%Ec) and mortality percentage (%Mt). The groups treated with 5,000 and 10,000 IJs/EU showed significant alterations in the PEP when compared to the control, while for the EP only the treatment with 10,000 IJs/EU produced any significant changes compared to the control. The MP did not differ significantly from the control in any of the treated groups. For the %Ec only the group treated with 10,000 IJs/EU differed significantly from the control. The greatest efficiency was obtained at the concentration of 10,000 IJs/EU, in which the mortality percentage was 13%. This demonstrates that the nematode utilized was not able to cause high mortality in engorged A. cajennense nymphs and agrees with other reports indicating that immature ticks are not highly susceptible to entomopathogenic nematodes.

Esse trabalho avaliou os efeitos in vitro de Steinernema glaseri sobre a biologia de ninfas ingurgitadas de Amblyomma cajennense. Os carrapatos foram expostos a juvenis infectantes (JI) de S. glaseri sob diferentes concentrações. Cada tratamento foi composto de 10 unidades experimentais (EU = repetições) em placas de Petri mantidas sob 27 ± 1º C e UR > 80 ± 10% por 72 horas. Em seguida, as ninfas foram transferidas para placas de Petri isentas de nematoides para observação dos seguintes parâmetros: período de pré-ecdise (PPE), período de ecdise (PE), período de muda (PM), porcentagem de ecdise (%Ec) e porcentagem de mortalidade (%Mt). Os grupos tratados com 5.000 e 10.000 JI/UE mostraram alterações significativas no PPE quando comparados com o controle, enquanto para o PE somente o tratamento com 10.000 JI/UE produziu mudanças significativas quando comparado com o controle. Não houve diferença significativa do PM em relação ao grupo controle em nenhum dos tratamentos e, para a %Ec, somente o grupo tratado com 10.000 JI/UE diferiu significativamente do controle. A maior eficiência foi obtida com a concentração de 10.000 JI/EU na qual a mortalidade foi de 13%. Isso demonstra que o nematoide utilizado não foi capaz de causar alta mortalidade em ninfas ingurgitadas de A. cajennese e concorda com outros relatos indicando que carrapatos imaturos não são altamente susceptíveis a nematoides entomopatogênicos.

Highly sensitive C-reactive protein and male gender are independently related to the severity of coronary disease in patients with metabolic syndrome and an acute coronary event

Patients with metabolic syndrome are at high-risk for development of atherosclerosis and cardiovascular events. The objective of this study was to examine the major determinants of coronary disease severity, including those coronary risk factors associated with metabolic syndrome, during the early period after an acute coronary episode. We tested the hypothesis that inflammatory markers, especially highly sensitive C-reactive protein (hsCRP), are related to coronary atherosclerosis, in addition to traditional coronary risk factors. Subjects of both genders aged 30 to 75 years (N = 116) were prospectively included if they had suffered a recent acute coronary syndrome (acute myocardial infarction or unstable angina pectoris requiring hospitalization) and if they had metabolic syndrome diagnosed according to the National Cholesterol Education Program/Adult Treatment Panel III. Patients were submitted to a coronary angiography and the burden of atherosclerosis was estimated by the Gensini score. The severity of coronary disease was correlated (Spearman’s or Pearson’s coefficient) with gender (r = 0.291, P = 0.008), age (r = 0.218, P = 0.048), hsCRP (r = 0.256, P = 0.020), ApoB/ApoA ratio (r = 0.233, P = 0.041), and carotid intima-media thickness (r = 0.236, P = 0.041). After multiple linear regression, only male gender (P = 0.046) and hsCRP (P = 0.012) remained independently associated with the Gensini score. In this high-risk population, male gender and high levels of hsCRP, two variables that can be easily obtained, were associated with more extensive coronary disease, identifying patients with the highest potential of developing new coronary events.

TNF-á and IL-6 immunohistochemistry in rat renal tissue experimentaly infected with Leptospira interrogans serovar Canicola

Leptospirosis is a public health problem worldwide and its etiology remains unclear. Its pathogenesis involves a complex interaction between host and infecting microorganism. The inflammatory reaction that controls the infection process also underscores many pathophysiological events occurring in leptospirosis. We investigated the presence of tumor necrosis factor-alfa (TNF-alfa) and interleukin-6 (IL-6) in renal tissues by immunohistochemical and histopathological examination in animals experimentally inoculated with Leptospira serovar Canicola. All the tests were carried out 2, 7, 14, 21 or 28 days after inoculation. Although TNF-alfa and IL-6 had been detected in tissues throughout the observation period, these cytokines appeared more intensely during the initial phase of infection. Therefore, both TNF-alfa and IL-6 were associated with the immunopathogenesis of leptospirosis. This profile suggests a high immunocellular response throughout the early infection stages followed by subsequent humoral response.

Fertility of male adult rats submitted to forced swimming stress

We investigated whether stress interferes with fertility during adulthood. Male Wistar rats (weighing 220 g in the beginning of the experiment) were forced to swim for 3 min in water at 32ºC daily for 15 days. Stress was assessed by the hot-plate test after the last stressing session. To assess fertility, control and stressed males (N = 15 per group) were mated with sexually mature normal females. Males were sacrificed after copulation. Stress caused by forced swimming was demonstrated by a significant increase in the latency of the pain response in the hot-plate test (14.6 ± 1.25 s for control males vs 26.0 ± 1.53 s for stressed males, P = 0.0004). No changes were observed in body weight, testicular weight, seminal vesicle weight, ventral prostate weight or gross histological features of the testes of stressed males. Similarly, no changes were observed in fertility rate, measured by counting live fetuses in the uterus of normal females mated with control and stressed males; no dead or incompletely developed fetuses were observed in the uterus of either group. In contrast, there was a statistically significant decrease in spermatid production demonstrated by histometric evaluation (154.96 ± 5.41 vs 127.02 ± 3.95 spermatids per tubular section for control and stressed rats, respectively, P = 0.001). These data demonstrate that 15 days of forced swimming stress applied to adult male rats did not impair fertility, but significantly decreased spermatid production. This suggests that the effect of stress on fertility should not be assessed before at least the time required for one cycle of spermatogenesis

Hormonal and metabolic study of a case of adiposis dolorosa (Dercum's disease)

A case of a 43-year-old nonobese woman with adiposis dolorosa (Dercum's disease) is reported. Muscle glucose uptake and oxidation before and after ingestion of 75 g of glucose were similar to control group values, although a greater insulin release(16,578 vs 6,242 ñ 1,136 uU/3 h) occurred simultaneously. In vitro studies of abdominal normal and painful subcutaneous adipose tissue of the patient revealed lower responsiveness to norepinephrine and lack of response to the antilipolytic effect of insulin in the painful adipose tissue (0.98 vs 1.43 uM FFA/106 cells at 5.0 uM of norepinephrine). The disease was not correlated with the HLA system and there were no alterations in hormonal secretion at the pituitary, adrenal, gonadal, and thyroid levels. These findings indicate the presence of peripheral insulin resistance in this patient with adiposis dolorosa

Association of specific histocompatibility antigens and acanthosis nigricans with insulin resistance

The association or interaction of histocompatibility antigens (HLA) and acanthosis nigricans with type A insulin resistance was studied in 13 patients (10 from family I, 2 from family II and an isolated case) for both sexes. HLA typing for the A, B, C and D antigens was performed by a standard microcytotoxicity test for all patients and for 100 normal controls from the same geographic area. The frequency of HLA B8 was 21% in the control group and 100% in patients with acanthosis nigricans. The frequency of HLA A1B8 was 15% in controls and 73% n acanthotic patients. All the members of family I presenting the association of a possibile insulin receptor defect (most likely provided by patient 2) with HLA B8 (provided by patient I) showed a more pronunced clinical and laboratory expression of insulin resistance. These data suggest that class I antigens of major histocompatibility complex (MHC), A1 and/or B8, may be involved in the pathogenesis of some forms of insulin resistance such as acanthosis nigricans (type A syndrome), possibly by a molecular interaction of the antigens with insulin receptors