2,3,7,8-Tetrachlorodibenzo-p-dioxin alters steroid secretion but does not affect cell viability and the incidence of apoptosis in porcine luteinised granulosa cells

The compound 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a by-product of human industrial activity, was found to affect ovarian steroidogenesis in animals, but the mechanism of its action is still unclear. The aims of the study were to examine the effect of TCDD on (1) progesterone (P4) and oestradiol (E2) production by granulosa cells isolated from medium (3–6 mm) and preovulatory (≥ 8 mm) porcine follicles, (2) the viability of the cells, and (3) the incidence of apoptosis. Porcine granulosa cells were cultured (48 h) with or without TCDD (100 pM, 100 nM). Steroid hormone concentrations in the medium were determined by radioimmunoassay. The viability of granulosa cells was tested spectrophotometrically (alamarBlue™ assay). Apoptosis was evaluated by flow cytometry using Annexin V and by TUNEL assay. The higher dose of TCDD (100 nM) significantly inhibited P4 and stimulated E2 production by luteinised granulosa cells isolated from medium follicles. The lower dose of TCDD (100 pM) significantly stimulated P4 and inhibited E2 secretion by the cells isolated from preovulatory follicles. None of the two TCDD doses affected cell viability or induced apoptosis in granulosa cells. In conclusion, TCDD directly affected steroid production by granulosa cells obtained from mature pigs, but the effect of TCDD was not due to its cytotoxicity

Comparative transcriptomic and proteomic signature of lung alveolar macrophages reveals the integrin CD11b as a regulatory hub during pneumococcal pneumonia infection

IntroductionStreptococcus pneumoniae is one of the main causes of community-acquired infections in the lung alveoli in children and the elderly. Alveolar macrophages (AM) patrol alveoli in homeostasis and under infectious conditions. However, the molecular adaptations of AM upon infections with Streptococcus pneumoniae are incompletely resolved.MethodsWe used a comparative transcriptomic and proteomic approach to provide novel insights into the cellular mechanism that changes the molecular signature of AM during lung infections. Using a tandem mass spectrometry approach to murine cell-sorted AM, we revealed significant proteomic changes upon lung infection with Streptococcus pneumoniae.ResultsAM showed a strong neutrophil-associated proteomic signature, such as expression of CD11b, MPO, neutrophil gelatinases, and elastases, which was associated with phagocytosis of recruited neutrophils. Transcriptomic analysis indicated intrinsic expression of CD11b by AM. Moreover, comparative transcriptomic and proteomic profiling identified CD11b as the central molecular hub in AM, which influenced neutrophil recruitment, activation, and migration.DiscussionIn conclusion, our study provides novel insights into the intrinsic molecular adaptations of AM upon lung infection with Streptococcus pneumoniae and reveals profound alterations critical for effective antimicrobial immunity

Microsatellite loci in the tetraploid spined loach, Cobitis biwae, and cross-species amplification in four related species

Fifteen microsatellite loci were identified in the tetraploid spined loach, Cobitis biwae (Teleostei: Cobitidae). Among these, 14 were polymorphic (5-31 alleles) and showed moderate to high cross-species amplification transferability in four related species, Cobitis matsubarai, Cobitis taenia, Misgurnus anguillicaudatus, and Misgurnus fossilis. The loci, described herein, will be useful for population genetics, phylogeny, parentage analysis, and detection of hybridization among Cobitis species

Analysis of testicular germ cell apoptosis in Cobitis loaches (Cobitidae, Teleostei) from diploid-tetraploid populations

Taxa of the genus Cobitis are distributed in Europe mainly in populations composed of individuals at different ploidy level. The species occur in Poland include the spined loach C. taenia and the Danubian loach C. elongatoides, triploid Cobitis females and tetraploid males and females. Previous morphological study has demonstrated abnormalities in spermatogenesis of the Cobitis tetraploid individuals associated with lack of sperm in their gonads. So far, however, the mechanism responsible for the gonadal sterility has not been explained. Over the past years, there is a growing evidence on incidence of apoptosis during fish spermatogenesis, however there is no data on the process of apoptosis in the male gonad of the natural polyploid. The aim of the study was to analyze apoptosis in the gonads of diploid and tetraploid Cobitis males. The expression of caspase-3 was analyzed by fluorescent immunohistochemistry using first rabbit, polyclonal anty-caspase-3 antibody (1:400) and second anti-rabbit antibody conjugated with Alexa Fluor 488 (1:2000). Number of TUNEL-positive cells was determined using fluorescence method based on the ability of terminal deoxynucleotidyl transferase (TdT) to incorporate fluorescein-labelled deoxyuridine triphosphate (dUTP) to the 3'-OH termini of the nuclear DNA strand breaks (TUNEL assay). Analysis of microscopic images showed no sperm in the testes of tetraploid Cobitis males. The highest expression of caspase-3 as well as the highest number of TUNEL-positive cells were demonstrated in the testes of the Danubian spined loach C. elongatoides compared to spined loach C. taenia and tetraploid males from diploid-polyploid population. The relatively low immunoexpression of caspase-3 and low number of TUNEL-positive cells do not unequivocally confirm that apoptosis disturb spermatogenesis in tetraploid males of Cobitis. Further studies are needed to explore the mechanisms behind such gonadal abnormality