Transglutaminase-mediated modifications of the rat sperm surface in vitro.

Two transglutaminase-mediated modifications of the rat epididymal spermatozoon surface were demonstrated in vitro. Transglutaminase was effective in promoting the binding of spermidine to the sperm. Moreover, the enzyme, by reacting with one of the major proteins secreted by the rat seminal vesicle epithelium, produced a modified form of the protein with a higher molecular weight and the capability of binding to the sperm cells. A specific physiological role for the enzyme, bringing about modifications of the rat sperm surface in the seminal fluid environment, is suggested.

Location of 64K collagen producer chondrocytes in developing chicken embryo tibiae.

The synthesis of a new low-molecular-weight collagen by cultured chicken embryo chondrocytes has been recently demonstrated (Capasso et al., Exp. Cell Res. 142:197-206, 1982; Gibson et al., J. Cell Biol. 93:767-774, 1982; Schmid and Conrad, J. Biol. Chem. 257:12444-12450, 1982). In this paper we report results on the location of chondrocytes synthesizing this new collagen (64K collagen) in the developing chicken embryo. The 64K collagen is synthesized in very large amounts by cells concentrated at the diaphysis of 9-day-old and at the epiphysis of 17-day-old embryo tibiae. These regions are characterized by a remodeling of the cartilage matrix leading to the replacement of the cartilage with bone tissue; therefore, this collagen appears to be a marker of a specific developmental stage of chondrocytes. The origin of cells competent for the synthesis of the 64K collagen is also discussed.

Block in the expression of differentiation markers of rat thyroid epithelial cells by transformation with Kirsten murine sarcoma virus.

Well-differentiated epithelial cells, derived from primary cultures of normal rat thyroid glands (T-79 cells), as well as a cloned cell line also derived from normal rat thyroid glands (FRT-L cells) were infected with Kirsten murine sarcoma virus carrying outer coat of the helper Kirsten murine leukemia virus. Infected T-79 and FRT-L cells changed morphologically and began to proliferate rapidly, suggesting malignant transformation by the virus. Both cell lines can support the replication of both transformation-competent and transformation-incompetent viruses such as murine or rat leukemia viruses. Infected T-79 and FRT-L cells had a high colony-forming efficiency (68 and 64%, respectively) when grown in agar and formed tumors when transplanted s.c. into syngeneic rats. These tumors morphologically resemble undifferentiated adenocarcinomas, thus showing that Kirsten sarcoma virus carrying the outer coat of the helper Kirsten murine leukemia virus is able to transform differentiated epithelial cells. Transformed T-79 and FRT-L cells, in contrast to uninfected cells, neither secrete thyroglobulin concentrate iodide, two biochemical markers of differentiated thyroid function. Thus, expression of the differentiated phenotype is blocked as a consequence of cell transformation. The system described may be useful in studying epithelial cell carcinogenesis in terms of regulated expression of differentiated functions.

From modeling to remodeling of upper airways: Centrality of hyaluronan (hyaluronic acid).

After traumatic events (accidental or surgical), the respiratory tract activates specific and prolix repairing mechanisms which tend to claw back the primitive differentiated state. The attempt of reactivation of the normal tissue functions is called 'remodeling' and its aim is to reinstate the modeling mechanisms that existed before the damaging event or the pathology's establishment. Endoscopic sinus surgery represents the gold standard treatment for inflammatory, malformative, benign, and, in selected cases, malignant diseases. The surgical technique is commonly described as minimally invasive as the nostrils are used as an access route and therefore does not leave any external scars. Currently, the surgical procedures, even though minimally invasive regarding the way in, are in fact widely destructive towards the surgical target. The healing process and re-epithelialization will depend on the amount of bony tissue that has been exposed and it will be important to stratify the different surgical typologies in order to foresee the increasing difficulty of mucosal healing process. As far as upper inflammatory diseases are concerned, recent studies demonstrated how intranasal hyaluronic acid can positively regulate mucosal glands secretion and modulate inflammatory response, being a useful tool for the improvement of remodeling after endoscopic sinus surgery. Acid has shown to be able to regulate mucosal glands secretion and modulate the inflammatory response.

Protective role of nuclear factor kappa B against nitric oxide-induced apoptosis in J774 macrophages.

We investigated the role of constitutive transcription factor nuclear factor kappaB (NF-kappaB) in nitric oxide (NO)-mediated apoptosis in J774 macrophages. Our results show that NF-kappaB is present in untreated J774 cells in a form constitutively active. Incubation of cells with sodium nitroprusside (SNP) and S-nitroso-glutathione (GSNO), two NO-generating compounds, caused: (a) inhibition of constitutive NF-kappaB/DNA binding activity; (b) decrease of cell viability; (c) DNA fragmentation; (d) ApopTag positivity. Pyrrolidine dithiocarbamate (PDTC) and N-alpha-para-tosyl-L-lysine chloromethyl ketone (TLCK), two inhibitors of NF-kappaB activation, showed the same effects of both NO-generating compounds. Furthermore, SNP and GSNO as well as PDTC and TLCK significantly increased the cytoplasmic level of IkappaBalpha. All together these results demonstrate that constitutive NF-kappaB protects J774 macrophages from NO-induced apoptosis. Moreover, these findings show, for the first time, that NO-generating compounds may induce apoptosis in J774 macrophages by down-regulating constitutive NF-kappaB/DNA binding activity and suggest a novel mechanism by which NO induces apoptosis.

Rat peritoneal immune response to carbon fibre reinforced epoxy composite implants.

The aim of this paper was to evaluate the histocompatibility of differently cured carbon fibre reinforced epoxy composites, studying their potential to induce an intolerance reaction in neighbouring tissues after peritoneal implantation in the rat. According to the microscopic and scanning electron microscope findings, the inductive capacity to generate connective tissue and cellular reaction was greatest in the partially cured material compared to the fully cured material. In addition, only the partially cured material implants appeared totally coated by macrophages at various stages of activation. The differences in the cellular reactions and scar tissue deposition in the interstices of these two composites are probably related to the chemical surface properties rather than to the structural characteristics of the materials.

Evidence that protease activated receptor 2 expression is enhanced in human coronary atherosclerotic lesions.

AIM: To investigate protease activated receptor 2 (PAR-2) expression in human coronary atherosclerotic lesions because PAR-2 is involved in the modulation of inflammatory events and vascular function. METHODS: An immunohistochemical analysis was performed on serial arterial sections, using the following antibodies: MDA2, a murine monoclonal antibody against malondialdehyde lysine epitopes of oxidised low density lipoprotein (oxLDL); HAM-56, a monoclonal antibody against human macrophages/foam cells; B5, a rabbit polyclonal antibody against PAR-2; and SAM11, a mouse monoclonal antibody against human PAR-2. Sections containing at least one lesion showing substantial immunostaining were counted as positive, and results were expressed as per cent of all sections of the same artery. RESULTS: PAR-2 expression was enhanced in human coronary atherosclerotic lesions. This phenomenon correlated with an increase in oxLDL epitopes in the coronary artery. CONCLUSION: This study shows for the first time that PAR-2 expression is enhanced in human coronary atherosclerotic lesions, and suggests that PAR-2 dependent cellular trafficking may be one of the regulatory signalling responses to vascular injury. Further pharmacological studies will establish whether modulation (and in which direction) of PAR-2 represents a possible therapeutic target for controlling the vascular response to injury.

Modulation of cytokine production in activated human monocytes by somatostatin.

The immunosuppressor effects of the widely distributed neuropeptide somatostatin were examined on purified peripheral blood human monocytes. Somatostatin, at concentrations thought to be physiologic (10(-10)-10(-7) M), regulated monocyte/macrophage responses to (LPS) stimulation, as reflected by interleukin production. In particular, somatostatin had direct inhibitory effects on TNF-alpha, IL-1 beta, and IL-6 secretion by LPS-activated monocytes, while the decrease on IL-8 synthesis was modulated mainly by the action of somatostatin on TNF-alpha and IL-1 beta. In fact, the addition of these two inflammatory cytokines to the monocyte culture medium was able to induce IL-8 expression, as demonstrated by mRNA analysis, also in presence of the neuropeptide. Although somatostatin affected IL-8 production in an indirect way, it suppressed directly the chemotactic response of neutrophils to IL-8. Finally, somatostatin downregulation of monocyte activation was confirmed by the decrease of HLA-DR expression on cell plasma membranes (52% versus 33%). Our results confirm that somatostatin exerts preferential effects on the suppression of immunoreactions by modulating cytokine production and activity.

A standardized procedure for using human corpus cavernosum strips to evaluate drug activity.

The main problem of using human corpus cavernosum (HCC) tissue to perform bioassay is linked to its limited availability further complicated by the heterogeneous source of the tissues used. Here, we show that gender reassignment is a reliable source of human tissue without major ethical problems. Indeed, the entire corpus cavernosum is obtained from the surgery procedure, which allows creating a standardized procedure to prepare HCC strip. In addition, human tissue, if kept in the fridge in the condition described, does not loose its ability to contract to phenylephrine (PE; alpha agonist), angiotensin II (AG II) and KCl up to 4 days. Furthermore, once contracted with PE, HCC relaxes to acetylcholine (endothelium-dependent mechanism); sodium nitroprusside (endothelium-independent mechanism); cromakalim (CRK), a K(ATP) channel opener; or alprostadil, a synthetic PGE2 (ALPR). In conclusion, we have standardized a procedure that allows the use of HCC strips to evaluate drug activity and/or to study pathophysiological mechanisms with an intact functional human tissue up to 4 days from the surgery procedure.

The structure and development of osteogenetic repair tissue according to Ilizarov Technique in man. Characterization of extracellular matrix.

Distraction osteogenesis by the method of Ilizarov has permitted the study of bone formation. This is an analysis of 64 human biopsies in patients who were undergoing tibial lengthening by the method of Ilizarov. After analysis by conventional and polarized light microscopy, four stages of bone formation were identified. Bone formation is of the direct type with no cartilaginous phase.

[End-to-side anastomosis made with a short circulatory interruption in the receiving vessel. Experimental model in the rat]. / Anastomose termino-latérale réalisée avec une brève interruption circulatoire du vaisseau receveur. Modèle expérimental chez le rat.

End to side microsurgical anastomoses between vessels of small calibre require interruption of blood flow of about 25-40 min. In terminal vascular beds this time may be incompatible with the cellular survival. The authors refer their experience in rats with a partly original experimental technique requiring less time of clamping the receiving vessel (about 3-4 min), as the main part of the anastomosis is performed between the wall of the donor vessel and the adventitia of the receiving vessel, without interruption of blood flow in this latter. Histological examinations, performed 21 days later, reveal a patency rate of 60%.

Peripheral eosinophil counts correlate with nasal eosinophil counts in patients with rhinitis

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Is there a correlation between osteoclastic activity and the action of calcitonin on macrophagic activity?

The formation of granulomas from a foreign body was studied by means of the subcutaneous implantation of cotton fragments in rats treated with salmon calcitonin or with just the diluent. The formation of granulomas was evaluated in optical microscopy, transmission and scanning electron microscopy (TEM and SEM, respectively). The cellular populations of the granuloma were immunophenotyped and flow analyzed in cytofluorometry (FACS = fluorescence-activated cell sorter). In the group treated with calcitonin, a lesser inflammatory effect of the cotton fragments was found compared to the group treated with only the diluent. These results indicate that, in addition to inhibiting bone resorption, salmon calcitonin has several important anti-inflammatory effects.

[Ultrastructure of human cerebral neoplasms in vitro: glioma]. / Ultrastruttura di neoplasie cerebrali umane in vitro: il glioma.

The authors describe the in vitro structural features of cellular populations derived from human gliomas. They report the structure of each population, and hypothesize on the correlation among these different cell populations. The findings of this study show some of the individual glioma cell lines to have characteristic structures.

Synthesis of seminal ribonuclease in isolated lobules of bull seminal vesicles.

A procedure is described for preparing and maintaining in culture isolated lobules of bovine seminal vesicles, consisting of glandular acini, surrounded by little connective tissue and with free access to the external medium, in which secreted material can be collected. After 48 h in culture, the isolated lobules appeared indistinguishable, by morphological and biochemical criteria, from freshly isolated lobules. After much longer culture times about one third of the glandular cells were still capable of effective protein synthesis. Studying the biosynthesis of seminal ribonuclease with preparations of isolated lobules we found that the enzyme was synthesized and secreted; only the fully amidated isoenzyme was synthesized and secreted, indicating that production of the selectively deamidated isoenzymic forms occurred after secretion, newly synthesized protein was rapidly exported, indicating that the high levels of enzyme previously reported for the seminal vesicle tissue were essentially due to its content of stored secretion.

[Ultrastructure of human cerebral neoplasms in vitro: medulloblastoma]. / Ultrastruttura di neoplasie cerebrali umane in vitro: medulloblastoma.

The fine structure of five lines of medulloblastoma cells in long-term cell cultures are described. The cell cultures were fixed in situ by the rapid exchange of the medium for the fixative solution consisting of 3% glutaraldhyde in 0.1 M Na cacodylate-HC1 buffer with 0.1 M sucrose (pH 7,2, total osmolality 510 mOsm, vehicle osmolality 300 mOsm). The cultures were subsequently postfixed in 1% OsO4 in water at room temperature and dehydrated in a graded series of ethanol solutions. In the second change of 100% ethanol they were stained with 2% uranyl acetate for 10 minutes. They described the structural characteristics of these cells in relation to the various phases of the culture, is similar to the well known of in vitro cellular embryonal systems (spongioblasts).

Mandibular lengthening by external distraction: an experimental study in the rabbit.

OBJECTIVE: To observe the development of bone at different times during the period of mandibular lengthening by external distraction using Ilizarov's transosseous osteosynthesis technique. MATERIALS AND METHODS: Fifteen rabbits, 2 to 3 kg in weight, were used for this experiment. The left side of the mandible was exposed and a corticotomy was performed with a water-cooled drill. After 12 hours, distraction was started and continued 1 mm per day for 2 weeks. Plain radiography and three-phase bone scan scintigraphy were performed on the 1st, 7th, and 14th postoperative days. The mandibles were then either removed for immediate histologic evaluation or after 2, 5, or 8 weeks of postdistraction fixation. RESULTS: Elongation of up to 11 mm was achieved. Radiographic and scintigraphic evaluation suggested a residual inflammation on the 7th day and definite ossification on the 14th day. Histologic observations of the distraction site showed a gradual change from an amorphous matrix to a fibrous matrix and, finally, an osseous-like tissue. CONCLUSION: Bone lengthening by gradual distraction is commonly used in tubular bones. To achieve mandibular lengthening it is necessary to resolve many problems. Morphologic, radiographic, scintigraphic, and histologic observations confirm that mandibular lengthening by external distraction is possible, with the formation of new bone tissue.

Intermediate filaments in a human medulloblastoma cell strain.

The fine structure of a human medulloblastoma cell strain characterized by the emission of two types of cytoplasmic processes was studied with particular regard to the cytoskeleton organization of the cell processes. A particular abundance of intermediate filaments (6-11 nm) was found in the primary processes. This study suggests that this abundance can strongly condition the cell shape of in vitro medulloblastoma cells and can represent a marker of this type of cells.