Intermediate-conductance Ca2+-activated K+ channels (IKCa1) regulate human prostate cancer cell proliferation through a close control of calcium entry.

Accumulating data point to K(+) channels as relevant players in controlling cell cycle progression and proliferation of human cancer cells, including prostate cancer (PCa) cells. However, the mechanism(s) by which K(+) channels control PCa cell proliferation remain illusive. In this study, using the techniques of molecular biology, biochemistry, electrophysiology and calcium imaging, we studied the expression and functionality of intermediate-conductance calcium-activated potassium channels (IK(Ca1)) in human PCa as well as their involvement in cell proliferation. We showed that IK(Ca1) mRNA and protein were preferentially expressed in human PCa tissues, and inhibition of the IK(Ca1) potassium channel suppressed PCa cell proliferation. The activation of IK(Ca1) hyperpolarizes membrane potential and, by promoting the driving force for calcium, induces calcium entry through TRPV6, a cation channel of the TRP (Transient Receptor Potential) family. Thus, the overexpression of the IK(Ca1) channel is likely to promote carcinogenesis in human prostate tissue.

Molecular typing of Pneumocystis jirovecii found in formalin-fixed paraffin-embedded lung tissue sections from sudden infant death victims.

Previous studies have provided histological evidence of an association between primary Pneumocystis infection and sudden infant death syndrome (SIDS). The aim of this work was to determine the species of clustered Pneumocystis organisms found in formalin-fixed paraffin-embedded (FFPE) lung tissue sections from Chilean sudden infant death (SID) victims. This approach needed first to optimize a DNA extraction method from such histological sections. For that purpose, the QIAamp DNA Isolation from Paraffin-Embedded Tissue method (Qiagen) was first tested on FFPE lung tissue sections of immunosuppressed Wistar rats inoculated with rat-derived PNEUMOCYSTIS: Successful DNA extraction was assessed by the amplification of a 346 bp fragment of the mitochondrial large subunit rRNA gene of the Pneumocystis species using a previously described PCR assay. PCR products were analysed by direct sequencing and sequences corresponding to Pneumocystis carinii were found in all the samples. This method was then applied to FFPE lung tissue sections from Chilean SID victims. Pneumocystis jirovecii was successfully identified in the three tested samples. In conclusion, an efficient protocol for isolating PCR-ready DNA from FFPE lung tissue sections was developed. It established that the Pneumocystis species found in the lungs of Chilean SID victims was P. jirovecii.

Interposition vein cuff and intimal hyperplasia: an experimental study.

OBJECTIVE: There is some evidence to suggest that prosthetic distal bypass graft patency can be improved, and the risk of intimal hyperplasia diminished, by interposing a distal vein cuff. We studied intimal remodeling in an end-to-side distal prosthetic anastomosis constructed with and without a vein cuff. METHODS: Twenty-four prosthetic bypasses were constructed with (N=12) or without (N=12) a distal vein cuff in 12 pigs. At 10 weeks, the 20 anastomoses and adjacent arteries from the surviving 10 pigs were studied by histology, immunohistochemistry and morphometry. RESULTS: Intimal hyperplasia was significantly less on all zones of the arterial floor and all suture zone of arteries anastomosed with a vein cuff than within arteries anastomosed without a vein cuff (0.11 versus 0.34; p=0.001 and 0.35 versus 1.19; p=0.0001, respectively). Intimal hyperplasia was also more prominent within the vein cuff than within the recipient artery, with or without a vein cuff (1.35 versus 0.38; p=0.0001). CONCLUSION: An interposition vein cuff at the distal anastomosis between a prosthesis and an artery alters the distribution of intimal hyperplasia. By acting as an expansion chamber where intimal hyperplasia can develop harmlessly, the vein cuff may protect the arterial anastomosis from stenosis.

Immunocompetent hosts as a reservoir of pneumocystis organisms: histological and rt-PCR data demonstrate active replication.

The present study was conducted to further examine recent data suggesting that pneumocystosis could be transmitted between patients and healthcare workers in the hospital environment, as has been proven with Pneumocystis-infected SCID mice and immunocompetent Balb/c mice. Using an experimental design (i.e., SCID-Balb/c mouse airborne transmission system), the present work found that healthy host-to-healthy host transmission of Pneumocystis organisms can occur, and that 'second' healthy contacts are able to transmit the infectious organisms to immunocompromised hosts. Further tests designed to explore the behavior of Pneumocystis organisms in the lungs of immunocompetent hosts were performed using histological and molecular approaches (e.g. testing the expression of both cyclin-dependent serine-threonine kinase and heat-shock 70 protein in Pneumocystis). The results showed Pneumocystis organisms were able to replicate in the lungs of immunocompetent hosts, which indicates these hosts are a reservoir for Pneumocystis spp.

Immunohistochemical contribution to the study of morphine metabolism in Calliphoridae larvae and implications in forensic entomotoxicology.

Morphine was detected by immunohistochemistry on sections of third stage larvae of Calliphora vomitoria (Diptera, Calliphoridae) reared on minced beef meat previously treated with morphine hydrochloride. The detection was performed with an avidin-biotin-peroxidase-complex method. Positive specimens showed specific staining of the haemolymph and a more intense immunoreaction in an area located at the limit between exocuticle and endocuticle. These results constitute an evidence of morphine accumulation inside the cuticle of Diptera larvae during their development. During the pupariation, the larval cuticle is transformed into the sclerotized puparium. This study consequently points out the possibilities of analyzing empty pupariae when suitable tissues or living necrophagous insects are absent.

In vivo experimental evaluation of skin remodeling by using an Er:Glass laser with contact cooling.

BACKGROUND AND OBJECTIVE: Selective dermal remodeling consists of inducing collagen tightening, neocollagen synthesis, or both, without damage to the overlying epidermis. This experimental study aimed to evaluate an Er:Glass laser emitting at 1.54 micrometer combined with contact cooling to target the upper dermis while protecting the epidermis. STUDY DESIGN/MATERIALS AND METHODS: Male hairless rats were used for the study. Different fluences (26-30 J/cm(2)) by using single 3-ms pulse irradiation or pulse train irradiation (1.1 J, 3 Hz) and different cooling temperatures (+5 degrees C, 0 degrees C, -5 degrees C) were screened with clinical examination and histologic evaluation at 1, 3, and 7 days after laser irradiation. RESULTS: The clinical effects were clearly dose and temperature cooling dependent. It seemed that single pulse irradiation led to epidermal whitening in most cases, whatever the cooling temperature. Conversely, pulse train irradiation showed reproducible epidermal preservation and confinement of the thermal damage into the dermis. New collagen synthesis was confirmed by a marked fibroblastic proliferation, detected in the lower dermis at day 3 and clearly seen in the upper dermis at day 7. CONCLUSION: This new laser seems to be a promising new tool for the treatment of skin laxity, solar elastosis, facial rhytides, and mild reduction of wrinkles.

Ultrastructural and molecular characterization of Pneumocystis carinii isolated from a rhesus monkey (Macaca mulatta).

High levels of heterogeneity have been observed among isolates of Pneumocystis carinii derived from different mammalian host species. We report the characterization of P. carinii isolated from a rhesus monkey (Macaca mulatta), which was immunosuppressed as a result of infection with a chimeric simian-human immunodeficiency virus (SHIVsbg). Histopathological examination showed evidence of severe P. carinii pneumonia with a large predominance of trophozoite forms. Alveolitis consisted of typical foamy, honeycomb exudate, with only a few alveolar macrophages. The lung inflammatory response was rather moderate without type-2 pneumocyte hyperplasia or collagenosis. P. carinii organisms were sometimes observed in the bronchiolar lumen. Ultrastructurally, macaque-derived P. carinii was more similar to human- or rabbit-derived parasites than to mouse-derived P. carinii. Molecular studies were carried out on the macaque-derived P. carinii DNA at two genetic loci: the genes encoding the mitochondrial large subunit ribosomal RNA (mt LSU rRNA) and the mitochondrial small subunit ribosomal RNA (mt SSU rRNA). Comparison of the DNA sequences with those from P. carinii isolated from eight other host species demonstrated that the macaque-derived P. carinii was genetically distinct at both loci, and was more closely related to human-derived P. carinii than to P. carinii derived from non-primate sources. We propose that macaque-derived P. carinii be named Pneumocystis carinii f.sp. macacae.

Morphological and ultrastructural methods for Pneumocystis.

Pneumocystis is a eukaryotic unicellular microorganism with marked fungal affinities. All known life cycle stages of this parasite were observed in the lung of mammals. The cystic forms of this microorganism may be observed microscopically by using stains with affinity for the components of their relatively thick cell wall. However, about 100 years ago they were observed for the first time thanks to panoptic stains which do not stain their cell wall. Methanol-Giemsa technique as well as Giemsa-like rapid stainings are often used to reveal vegetative or cystic forms of this parasite on air dried smears of clinical or experimental samples. For many years, hypotheses on its life cycle, which remains unknown, were based on transmission electron microscopy (TEM) studies. However, only for the last years progresses in the quality of fixation for TEM led to a better understanding of the Pneumocystis cell structure. In this chapter, strategies to reveal Pneumocystis organisms in clinical or experimental specimens by using light microscopy, as well as techniques allowing a good preparation of parasitic samples for TEM, are given and shortly discussed.