Macrophage Migration Inhibitory Factor contributes to drive phenotypic and functional macrophages activation in response to Toxoplasma gondii infection.

Cytokines are small molecules secreted by numerous cells. Macrophage Migration Inhibitory Factor (MIF) is a cytokine initially described due to its function of inhibiting random macrophage migration. Currently, new functions have been described for MIF, such as stimulating inflammatory functions in response to infections by microorganisms including, Toxoplasma gondii. However, the primordial MIF function related to macrophages has been little addressed. The main purpose of the study was to recapitulate MIF function on macrophages in response to T. gondii infection. To achieve this goal, peritoneal macrophages were collected from C57BL/6WT and Mif1-/- mice after recruitment with thioglycolate. Macrophages were cultured, treated with 4-Iodo-6-phenylpyrimidine (4-IPP), and infected or not by T. gondii for 24 h. Following this, the culture supernatant was collected for cytokine, urea and nitrite analysis. In addition, macrophages were evaluated for phagocytic activity and T. gondii proliferation rates. Results demonstrated that T. gondii infection triggered an increase in MIF production in the WT group as well as an increase in the secretion of IL-10, TNF, IFN-γ, IL-6 and IL-17 in the WT and Mif1-/- macrophages. Regarding the comparison between groups, it was detected that Mif1-/- macrophages secreted more IL-10 compared to WT. On the other hand, the WT macrophages produced greater amounts of TNF, IFN-γ, IL-6 and IL-17. Urea production was more pronounced in Mif1-/- macrophages while nitrite production was higher in WT macrophages. T. gondii showed a greater ability to proliferate in Mif1-/- macrophages and these cells also presented enhanced phagocytic activity. In conclusion, T. gondii infection induces macrophage activation inciting cytokine production. In presence of MIF, T. gondii infected macrophages produce pro-inflammatory cytokines compatible with the M1 activation profile. MIF absence caused a dramatic reduction in pro-inflammatory cytokines that are balanced by increased levels of urea and anti-inflammatory cytokines. These macrophages presented increased phagocytic capacity and shared features activation with the M2 profile.

Effects of Rooibos (Aspalathus linearis) on the Testicular Parenchyma of Adult BALB/C Mice Submitted to a Hypercaloric Diet

Oxidative stress due to obesity plays a detrimental role in the testicular microenvironment and sperm parameters. We explored the impact of a hypercaloric diet in male BALB/c mice as a condition to trigger damage to the spermatogenic process and the antioxidant effect of Aspalathus linearis as well. We used a hypercaloric diet in animals divided into 3 groups: Control, Hypercaloric diet control (HC) and Hypercaloric diet and Rooibos infusion (HCR). Morphometric parameters, enzyme dosages, cell viability, and tubular histopathology were evaluated. Body weight increased in HCR animals at weeks 3, 4, and 8. We found a reduction in seminiferous epithelium height, with an increase in the tubular diameter of the HCR group. Catalase levels were lower in HC and HCR, while carbonyl protein was decreased in HC. We estimate that it induces oxidative stress (OS) capable of affecting the seminiferous epithelium and that the infusion of A. linearis does not demonstrate a potential benefit in cell preservation.

Doxycycline hyclate may damage testicular germinal epithelium in Wistar rats

Broad-spectrum antimicrobial doxycycline acts as an inhibitor of protein synthesis and it is widely used in the clinical treatment of various infections by microorganisms that are sensitive to the drug, as well as in animal feed. Its liposolubility guarantees its high tissue bioavailability, being associated with several biochemical changes in the organism and potentially adverse effects on reproduction. This study aims to evaluate the effects of the action of doxycycline on spermatogenesis to provide a complete analysis of the tubular and interstitial compartments and to identify possible changes in the testicular parenchyma. Adult male Wistar rats were divided into three groups: one control (water), and two treated with doxycycline at the doses of 10mg/kg and 30mg/kg, for 30 days. After euthanasia and sample processing, the following parameters were evaluated: a) tubular diameter and height of the seminiferous epithelium; b) volumetric proportions (%) and volumes (mL) of the components of the testicular parenchyma; c) counting testicular germ cell populations; d) evaluation of cell viability. The results of the comparative evaluation between the experimental groups demonstrated a significant increase in the diameter and area of the tubular lumen and a reduction in the count of spermatogonia in the experimental group that received doxycycline hyclate at a dose of 30mg/kg. In the same experimental group, an increase in the overall yield of spermatogenesis was found as a consequence of the increase in the mitotic index.

Comparison of the spermatogenic process in three different mice strains: Swiss, Balb/C and C57BL/6

Many studies have been trying to establish standard protocols for animal experimentation, especially for animal species or strains, to master research variables with high precision. The main mouse strains used in the field of the biology of reproduction are Swiss, Balb/c, and C57BL/6. Since some of the strains show reproduction limitations, such as the size of the litter, the present study aimed to compare their spermatogenic processes to verify differences regarding the testicular parenchyma and germ cell populations, which could explain low offspring production. In addition, the present study provides additional information concerning the testicular parenchyma of such strains, which consequently would help researchers to choose the most suitable strain for reproductive studies. Six adult male mice were used for each of the strains. After euthanasia, the testes were weighed, fixated with Karnovsky fixative, embedded in methacrylate, sectioned, and stained with toluidine blue/sodium borate 1%. Morphometrical analyses from the testicular parenchyma (seminiferous tubules and interstitium) were made using the software ImageJ. Germ and Sertoli cells populations were counted in seminiferous tubules cross-sections at stage I of the seminiferous epithelium cycle. The lowest body and testicular weights were observed in C57BL/6 animals, followed by Balb/c and Swiss, however, the relative testes, parenchyma, and albuginea weights were significantly lower only in C57BL/6. Despite the seminiferous tubules and seminiferous epithelium proportions were lower in Swiss animals, their relative amount related to the body weight was the same among strains. The total number of germ cells was higher in Swiss animals, reflecting higher spermatogenic yield and daily sperm production. Due to the lower relative number of Sertoli cells, the Swiss animals showed the highest Sertoli cell index and support capacity. On the other hand, the lowest pathological indexes regarding the germ cells were observed in Balb/c animals, followed by Swiss and C57BL/6. In the interstitium, the proportion of blood vessels was lower in Swiss mice, while the lymphatic cell proportion was lower in C57BL/6 animals. Moreover, the highest proportions of Leydig cells and macrophages were noticed in Swiss mice, which may indicate increased testosterone levels. Altogether, such observations must be taken into account when choosing any of the studied strains for reproduction studies.

Pfaffia glomerata hydroalcoholic extract stimulates penile tissue in adult Swiss mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Roots of Pfaffia glomerata are used in traditional medicine as aphrodisiacs and sexual stimulants. AIM OF THE STUDY: The aim of this study was to evaluate the action of the hydroalcoholic extract from the roots of Pfaffia glomerata on the Leydig cells, cavernous bodies and other penile constituents, as well as on serum testosterone and 17ß-estradiol levels of adult mice. MATERIALS AND METHODS: Mature male Swiss mice were divided into 6 groups: control (water), sildenafil citrate, 3 groups receiving daily doses of P. glomerata extract (100, 200 and 400 mg/kg) and one group receiving intermittent doses of P. glomerata (200 mg/kg/3-3d). RESULTS: The proportions of blood vessels, lymphatic space and estradiol levels were increased. On the other hand, reduction of testosterone levels due to Leydig cells death was observed. As for penile parameters, volumetric proportions of cavernous bodies, collagen and nitric oxide were increased, while smooth muscle content was decreased. CONCLUSIONS: Despite that the long term intake of P. glomerata extract was related to a stimulant action, reduction on Leydig cell viability induced decreased testosterone production.

Hydroalcoholic extract of Pfaffia glomerata alters the organization of the seminiferous tubules by modulating the oxidative state and the microstructural reorganization of the mice testes.

ETHNOPHARMACOLOGICAL USAGES: Leaves and roots of Pfaffia glomerata areused as aphrodisiacs, tranquilizers and antirheumatics. Due to the lack of experimental scientific data, studies are necessary to identify its medicinal properties. AIMS: The present study aimed to evaluate the effect of hydroalcoholic root extract of P. glomerata (Brazilian ginseng extract - BGE) on testicular parenchyma, and evaluate possible harmful effects through testicular oxidative stress analysis. MATERIALS AND METHODS: Adult mice were divided into 6 groups: control (water), sildenafil citrate, BGE (100, 200 and 400 mg/kg/day), and BGE (200 mg/kg every three days). RESULTS: The treatment reduced the volumetric proportions of seminiferous tubules and epithelium, the number of Sertoli cells, and increased hydrogen peroxide levels, without affecting sperm production. It also caused cell death and changes in the frequency of stages of the seminiferous epithelium cycles. The 100 mg/kg dose responds in a similar way to sildenafil citrate, promoting changes in the gonadal structure, but with efficient response to contain the damage. CONCLUSIONS: Doses of 200 mg/kg, continuous or discontinuous, induced an increase in testicular nitric oxide, as well as sildenafil citrate, showing be efficient as aphrodisiac, but promotes cell death regardless of the form of administration.

Stereological analyses of the annual variation of captive bullfrogadult testes (Lithobates catesbeianus, Shaw 1802)

The occurrence of seasonality in bullfrog (Lithobates catesbeianus) spermatogenesis was analyzed by quantifying germ cell populations throughout the year, which were divided into bimesters, from November 2001 to October 2002. Body weight, liver weight, fat body weight and gonadal weight were taken and bimonthly grouped. Using these data, the gonadosomatic, hepatosomatic, and liposomatic indexes were calculated. All animals were kept in the same cement cages in a frog farm, and sampled every month. The cages were underthe influence of climate variables changes, such as photoperiod, temperature, rainfall and relative humidity.Body weight was higher during Spring and Summer, and a significant reduction has occurred between Julyand August. The highest gonadosomatic index values were registered from May to October, coinciding withthe period of gonadal preparation for reproduction. Also, the reproduction of bullfrog is directly related toenvironmental factors, mainly due to Summer and Winter humidity variations.

Reproductive biology of Oligosarcus argenteus (Gunther, 1864) adult males and description of the gonadal maturation stages

Oligosarcus argenteus belongs to the Acestrorhynchinae subfamily, being restricted to South America, and found in several Brazilian hydrographic basins, in lotic and lentic environments, where they are able to reproduce. With the purpose of studying the reproductive biology of the males from this species, many morphological parameters were analyzed during a 24 month period, as well as characterizing the different testicular maturation stages. A maturity scale, with three stages (I - Initial Maturing, II - Intermediate Maturing, III - Final Maturing) was proposed for the adult males of Oligosarcus argenteus. The reproductive period was established by the bimonthly frequency of spermatogenesis and by the gonadal maturation stages.

Oligosarcus argenteus é uma espécie pertencente à subfamília Acestrorhynchinae, restrita à América do Sul, sendo comumente encontrada nas várias bacias hidrográficas brasileiras, em ambientes lóticos e lênticos, onde se reproduzem. Com o objetivo de estudar a biologia reprodutiva de machos desta espécie, vários parâmetros morfológicos foram analisados durante um período de 24 meses, assim como foram caracterizados os diferentes estádios de maturação testicular. Estes foram descritos, considerando-se exemplares adultos, em: Estádio I - Maturação Inicial, Estádio II - Maturação Intermediária e Estádio III - Maturação Final. Pela freqüência bimestral das células germinativas e dos estádios de maturação gonadal, a época de reprodução foi determinada.