Functional activity of varicose human great saphenous vein at different sites.

Experiments on isolated preparations of varicose human great saphenous vein revealed different sensitivity of the distal and proximal portions to P2X receptor agonist α,ß-methylene-ATP, but not to norepinephrine and histamine. It is suggested that restructuring of the P2 receptor system plays an important role in the pathogenesis of varicose veins, which affects the trunk of the great saphenous vein in varying degrees.

Are mesh anchoring sutures necessary in ventral hernioplasty? Multicenter study.

BACKGROUND: Avoiding mesh fixation to the surrounding tissue in ventral hernioplasty would simplify the operation, decrease the time of the procedure, and decrease the risk of suture-related complications. METHODS: Four hospitals included 111 patients according to the common protocol for prospective clinical evaluation of sutureless ventral hernioplasty. Surgical technique involves placement of the polypropylene mesh with flat-shape memory in either the retromuscular or preperitoneal space without suture anchoring. RESULTS: Local complication rate was low (12.6%, 14 patients), postoperative pain measured according to the visual analogue scale was minimal (mean 4, range 1-8). Three recurrences (3%) were recorded. Mild scar discomfort, which did not require treatment nor limit physical activity, was recorded in 28 (25%), 18 (17%), and 11 (14%) patients at 6-month, 1- and 2-year follow-up, respectively. CONCLUSIONS: Results of the study suggest that the sutureless sublay technique is safe and effective in the treatment of ventral abdominal hernia, especially in small and medium defects.

RXRalpha regulates the pregnancy-specific glycoprotein 5 gene transcription through a functional retinoic acid responsive element.

Human pregnancy-specific glycoproteins (PSG) are major placental polypeptides encoded by eleven highly conserved genes expressed by the syncytiotrophoblast. The minimal promoter region of all PSG genes contains a putative Retinoic Acid Responsive Element (RARE) though the ability of retinoids to regulate PSG gene expression has not been established. Retinoid signaling pathway plays a key role for overall placenta biology and is essential for trophoblast differentiation. In this work, we investigated the participation of the RARE motif in the regulation of PSG5 gene transcription by retinoic acid and its receptors. The minimal promoter region of PSG5 gene was activated by RXRalpha but not by RARalpha, in a ligand-dependent manner. The RARE sequence of PSG5 gene promoter was recognized by endogenous RXRalpha present in placental nuclear extracts as well as by RXRalpha either over expressed in cultured non-placental cells or in vitro translated. Mutations at specific nucleotides within the RARE motif abrogated both RXRalpha DNA binding and transcriptional activation of PSG5 promoter mediated by RXRalpha. Moreover, endogenous PSG expression was significantly induced in trophoblast-derived Jeg-3 cells upon 9-cis retinoic acid treatment. Interestingly, the induction level was higher following methotrexate-induced differentiation of Jeg-3 cells to syncytiotrophoblast-like structures. Altogether, these data provide the first evidences demonstrating that transcriptional activity of PSG5 gene is responsive to an external signal involving the retinoids-RXRalpha axis through a conserved RARE motif shared by all PSG gene family members.

Rating the severity of the medical consequences of drug-induced liver injury.

Risk analysis in drug development aims to allow for clear decisions showing whether or not the benefit of an intervention outweighs the risk. One of the difficulties in doing this in a repeatable and clear way is the problem of comparing different adverse events, as seriousness is often subjective. Using drug-induced liver injury as our model, we show that clinical, laboratory, and histological manifestations of liver reactions can be ranked by experienced hepatologists and these rankings can be used to rank the consequences of drug-induced side effects as a continuum. This risk ranked information could be transformed to standardized scores (z score) and the risks displayed by standard techniques; adverse events can then be compared with effects from other drugs and possibly with the consequences due to untreated disease or natural occurrences. As a risk is a function of both the seriousness of the event and the probability of its occurrence, risk can therefore be displayed in terms of probability and hazard to further ease communication. We propose that risk management of drugs in development would be improved, especially in terms of risk communication, if the hazard were ranked by means of a common scale and displayed in graphic form against the likelihood of occurrence.

Use of antibacterial agents in renal failure.

This article reviews the pharmacokinetics of antibacterial agents in patients with normal and decreased renal function. The concepts of volume and distribution, rate of elimination, loading and maintenance doses, and therapeutic drug monitoring are delineated. Special reference is made to the intermittent dosing of cefazolin with hemodialysis. Newer, as well as traditional methods of extracorporeal circulation and the resultant changes in antibacterial agent pharmacodynamics are discussed.

Use of antibacterial agents in renal failure.

This article provides information on the pharmacokinetics of antibacterial agents in patients with normal renal function and those with impaired renal function. Specific discussion includes the use of serum levels, dosage adjustments in dialysis, new strategies for cefazolin dosages in dialysis patients, and antibiotic toxicity in renal failure, and tabular data is presented for determining appropriate dosages for varying degrees of renal failure.

The Krüppel-like core promoter binding protein gene is primarily expressed in placenta during mouse development.

The human core promoter binding protein (hCPBP) has been identified as a DNA-binding protein involved in the regulation of TATA box-less genes like those encoding the pregnancy-specific glycoproteins. Structurally, hCPBP contains three zinc fingers in the C-terminal domain, which is highly conserved in a number of proteins that constitute the Krüppel-like family of transcription factors. In the present work, we report the molecular cloning of the mouse CPBP (mCPBP) and its expression pattern during development as well as in adult tissues. The mouse cDNA encodes a protein of 283 amino acids that share 94.4% of identity with the hCPBP. The highest level of mCPBP transcript was detected in placenta, and its expression was lower in total embryos and in adult tissues. We also show by in situ hybridization that during embryonic development the mCPBP gene is mainly expressed in extra-embryonic structures throughout gestation; essentially no specific expression was detected in embryonic tissues. Our data demonstrate that CPBP transcript is enriched in the trophoblastic tissue and strongly suggest that its encoded polypeptide regulates target genes involved in placental development and pregnancy maintenance.

Time course of biochemical and immunohistological alterations during experimental allergic encephalomyelitis.

A comprehensive biochemical, immunological and histological study was undertaken during different stages of experimental allergic encephalomyelitis (EAE). Wistar rats with EAE induced by sensitization with bovine myelin showed a maximum decrease of body weight 14-16 days post-inoculation (dpi), coincident with the appearance of the paralysis symptom (acute period). Quantitation of some brain components indicated a temporal dissociation among the alterations observed. The higher diminution of myelin basic protein (MBP) occurred at 6 dpi and then increased to reach 21 dpi, a normal value. Also, the activity of 2',3'-cyclic nucleotide 3'-phosphohydrolase was reduced by 40% with respect to control animals only at 6 dpi. The total lipid content was normal; however, among the individual lipids, sulfatides were principally degraded during the acute stage but the amount of cerebrosides was decreased during the recovery period (29-40 dpi). Free cholesterol was similar in both groups of animals, whereas cholesterol esters were detected in EAE animals from 14 to 40 dpi. Central nervous system meningeal and parenchymal infiltration with mononuclear cells was recognized principally at 14 dpi, but some of cells were still present at 40 dpi. Deposits of immunoglobulins in the infiltrated regions as well as in spinal cord motor neurons were observed among 14-29 dpi. Total circulating antibodies to MBP began to increase at 14 dpi, reaching a plateau at 21 dpi and then maintaining this value until 40 dpi. However, the population of anti-MBP antibodies that also recognizes the neuronal protein synapsin was only present at 14 dpi. The present results suggest that the neurological symptoms can be related to some early changes in the myelin membrane followed by alterations involving neuronal structures. The existence of immunological factors against some epitopes in MBP that also recognize a synaptosomal protein might account, at least in part, for the axonal damage and disruption of the normal interneuronal activity in EAE and lead together with the alterations in some specific myelin constituents and the concomitant CNS inflammatory process to the observed hindlimb paralysis.

Factor analysis of proficient esophageal speech: toward a multidimensional model.

This study identified acoustic patterns in the speech samples of 26 esophageal speakers judged by experienced listeners to be highly proficient and intelligible. Tape-recorded readings were acoustically analyzed in terms of frequency, intensity, and duration variables. Application of two multidimensional statistical procedures, factor analysis and cluster analysis, revealed four distinctive acoustic profiles that captured all 26 subjects. The multidimensional model derived from these profiles maintains important individual differences in alaryngeal speech style.

Chimeric B72.3 mouse/human (IgG1) antibody directs the lysis of tumor cells by lymphokine-activated killer cells.

Chimeric mouse/human B72.3 (cB72.3) antibodies having a human IgG1 (gamma 1) or IgG4 (gamma 4) constant region were compared to the native murine IgG1 B72.3 (nB72.3) monoclonal antibody (mAb) for their ability to participate with human effector cells in antibody-dependent cellular cytotoxicity (ADCC). Because the TAG-72 antigen recognized by B72.3 is poorly expressed on tissue-cultured tumor cell lines, the xenografted OVCAR-3 human ovarian carcinoma ascites was used as a cytotoxicity target. The lytic activity of the cB72.3(gamma 1) mAb with peripheral blood lymphocytes was 1.5- to 50-fold greater than that of the nB72.3 mAb and usually the cB72.3(gamma 4) mAb. However, lymphocytes from some donors had similar ADCC activity with either the cB72.3(gamma 1) or cB72.3(gamma 4) mAb. The cB72.3(gamma 1) and the murine anti-colon carcinoma CO17-1A mAb had comparable activity in mediating ADCC against the OVCAR-3 tumor. Exposure of lymphoid cells to interleukin-2 (IL-2) (100-500 U/ml) for 24 h to generate lymphokine-activated killer (LAK) cells augmented ADCC mediated by the cB72.3(gamma 1) mAb 2- to 22-fold. By contrast, LAK cells from most donors expressed weak non-specific cytotoxicity against OVCAR-3 ascites tumor cells. The cB72.3(gamma 1), and to a lesser extent, the cB72.3(gamma 4) chimera also participated with monocytes in mediating ADCC, but the antibody-dependent lytic potency of monocytic effectors was much weaker than that of IL-2-activated lymphoid cells. These studies show that the cB72.3(gamma 1) mAb has appreciable ADCC-mediating properties, suggesting a potential role for its incorporation into treatment strategies utilizing adoptive killer cell and/or lymphokine therapy.

Purification and identification of two serine class proteinases from dog mast biochemically and immunologically similar to human proteinases tryptase and chymase.

Serine class proteinases with trypsin-like and chymotrypsin-like specificity were purified from dog mastocytoma tissue. An antiserum was produced against the chymotrypsin-like proteinase. The antiserum reacted with mast cells in skin sections prepared from normal dogs consistent with the proteinase being a mast cell constituent. The antiserum also cross-reacted with the major chymotrypsin-like proteinase isolated from normal dog skin and partially cross-reacted with human skin chymase. No cross-reaction was detected with rat chymase. The trypsin-like proteinase from dog mastocytoma tissue was similar to tryptase isolated from human skin. It had a similar subunit structure, was not inhibited by many protein proteolytic enzyme inhibitors, bound to heparin, and reacted strongly with antiserum against human tryptase. Antiserum against human tryptase also reacted with mast cells in skin sections prepared from normal dog skin. No immunocytochemical labeling of rat skin mast cells was observed with anti-human tryptase. These studies establish the presence of a trypsin-like and chymotrypsin-like proteinase in dog skin mast cells and provide immunological evidence which suggests that both proteinases are more closely related to human than rat mast cell proteinases. These immunological and biochemical relationships are important when comparing the roles of these proteinases in different animals.

Studies of serum-free culture medium in the generation of lymphokine activated killer cells.

Lymphokine activated killer (LAK) cells administered in conjunction with recombinant interleukin-2 can mediate the regression of metastatic tumor in some patients with advanced cancer. In these trials LAK cells were activated in medium containing 2% human type A or AB serum. We have found three commercially available, serum-free culture media which allow development of in vitro LAK activity by human peripheral blood lymphocytes. They are AIMV (Gibco), MASF-3 (Whitaker-MA Bioproducts) and HB-104 (Dupont). If 2-mercaptoethanol was added to these culture media they were also capable of generating murine LAK cells which were effective in reducing pulmonary metastases in the murine MCA-106 model. Although LAK cells generated in these media have not been tested in humans yet, potentially they could provide a safe, unlimited and less expensive source of culture fluid for generating the large numbers of LAK cells needed for human clinical trials.

Development of an automated closed system for generation of human lymphokine-activated killer (LAK) cells for use in adoptive immunotherapy.

Immunotherapy utilizing the adoptive transfer of lymphokine-activated killer (LAK) cells in conjunction with recombinant interleukin-2 (IL-2) can mediate tumor regression in some patients with advanced cancer. The activation of large numbers of LAK cells was performed in roller bottles in a research laboratory setting and required meticulous aseptic technique, at least one skilled technician per patient and one laminar flow hood per patient. To reduce the complexity and expense of LAK cell generation for human immunotherapy trials we have developed a closed-system automated procedure using a continuous flow blood cell separator. PBL were obtained by standard apheresis techniques. Platelets and plasma were elutriated using countercentrifugal flow of saline in the cell separator machine. The washed PBL were underlaid with Ficoll-Hypaque (FH) in the original separation bag. Lymphocytes were then flushed into a collection bag where they were concentrated and washed with 2 liters of saline. Mean recovery from the automated FH technique was 54.6 +/- 4.3% compared to 62.3 +/- 4.0% using manual methods in 50 ml tubes (P greater than 0.05). Cells were diluted in the collection bag with RPMI 1640 +/- 2% human AB serum and could be dispensed in an automated fashion to polyolefin bags via a sample port with 1000-1500 U/ml IL-2. After 3-4 days of culture in 5% CO2 at 37 degrees C, activated cells from the bags were harvested and washed in a closed system using the continuous flow cell separator. Cell yield from the harvest was 79.2 +/- 5.4% in the automated system compared to 64.9 +/- 5.0% in the standard procedure using manual harvest of roller bottles (P less than 0.01). Lytic capacity of the cells against fresh human tumor in a 4 h 51Cr release assay was equivalent in cells processed either by the automated or the conventional manual method. The advantages of a closed system include decreased potential for microbial contamination and reduced labor and capital equipment costs. This technique may be easily adapted for use with other cell collection and culture systems.

Identification of a chymotrypsin-like proteinase in human mast cells.

An antiserum was produced against a chymotryptic proteinase purified from human skin. The antiserum did not cross-react with human leukocyte cathepsin G and elastase, rat mast cell proteinase I, and human skin tryptase. Indirect immunofluorescent staining of frozen skin sections to localize the proteinase showed cytoplasmic staining of cells scattered about the papillary dermis and around blood vessels and appendages. Restaining these sections with toluidine blue revealed that the fluorescently stained cells contained metachromatically staining granules, the major distinguishing feature of mast cells. A similar correlation was found in lung tissue. Ultrastructural studies employing the ferritin bridge technique to immunologically identify the proteinase additionally localized the proteinase to mast cell granules. Biochemical and immunochemical characterization of chymotryptic activity solubilized from isolated human lung mast cells identified a chymotryptic proteinase that may be identical to the skin chymotryptic proteinase. These studies establish that human skin mast cells contain a chymotrypsin-like proteinase that is a granule constituent and provide evidence that indicates a comparable proteinase is also present in lung mast cells.