Fibronectin initiates fibrin production by rabbit corneal endothelial cells in vitro

Confluent rabbit corneal endothelial cells incubated in the absence of serum do not produce fibrinogen. When exogenous fibronectin is added to these cultures, fibrinogen production is observed. Fibronectin concentrations stimulate fibrinogen synthesis by endothelial cells in a dose-response fashion. This direct interaction of fibronectin and fibrinogen may be important in both wound healing processes and pathological states.     

Collagen crosslinking and cartilage glycosaminoglycan composition in normal and scoliotic chickens

The amounts of lysine-derived crosslinks in collagens from tendon, cartilage, intervertebral disc, and bone and changes in the composition of sternal cartilage glycosaminoglycans were estimated in two lines of chickens, a control-isogenic line and a line that develops scoliosis. In the scoliotic line, scoliosis first appears at 3-4 weeks and progressively increases in severity and incidence so that 90% of the birds express the lesion by week 10. We have reported previously that cartilage, tendon, and bone collagens from scoliotic birds are more soluble than corresponding collagens from normal birds. Herein, collagen crosslinking and altered proteoglycan metabolism are examined as possible mechanisms for the differences in collagen solubility. At 1 week of age there were fewer reducible crosslinking amino acids (hydroxylysinonorleucine, dihydroxylysinonorleucine, and lysinonorleucine) in collagens from sternal cartilage and tendon in the scoliotic line than in the isogenic line. However, by week 3 and at weeks 5 or 7 values were similar in both groups. The amounts of hydroxypyridinium in vertebral bone and intervertebral disc collagen were also similar in both groups of birds. Consequently, differences in collagen crosslinking do not appear to be a persistent developmental defect underlying the expression of scoliosis in the model. However, differences were observed in cartilage proteoglycans and glycosaminoglycans from the scoliotic line that were not present in cartilage from the isogenic line. The average molecular weight of the uronide-containing glycosaminoglycans was 30% less in the scoliotic line than in the isogenic line, i.e., 12,000 compared to 18,000. The size distribution of cartilage proteoglycans from the scoliotic line also differed from that of proteoglycans from the isogenic line.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of semen fractionation and dilution ratio on equine spermatozoal motility parameters

Two experiments were conducted to examine the effects of semen fractionation and dilution ratio on motility parameters of stallion spermatozoa. In Experiment 1, three ejaculates from each of three stallions were divided into sperm-rich (SR) and sperm-poor (SP) fractions to determine the difference in sperm concentration. Mean sperm concentration in SR fractions (349.5 x 10(6)/ml) was greater (P < 0.001) than that of SP fractions (96.9 x 10(6)/ml). In Experiment 2, three ejaculates from each of two stallions were divided into SR and SP fractions. Fifty percent of the original volume of SR fractions was combined with 50% of the original volume of SP fractions for each ejaculate to represent total ejaculates. SR and total ejaculates were diluted with skim milk-glucose semen extender as follows: 1) no dilution, or dilution to 2) 100 x 10(6)sperm/ml, 3) 50 x 10(6)sperm/ml, or 4) 25 x 10(6)sperm/ml. Semen samples were evaluated at 0.5, 3, 6, 12, and 24 h postejaculation (25 degrees C storage temperature) for percentages of total spermatozoal motility (TSM) and progressive spermatozoal motility (PSM). Mean TSM was greater (P < 0.05) in SR ejaculates than total ejaculates at 12 and 24 h postejaculation. Mean TSM of undiluted semen was lower (P < 0.05) than other dilution ratios over all periods. Mean TSM was greater (P < 0.05) at a 25 x 10(6)sperm/ml dilution ratio than a 50 x 10(6)sperm/ml dilution ratio at 12 and 24 h postejaculation, and greater (P < 0.05) than a 100 x 10(6)sperm/ml dilution ratio from 3 to 24 h postejaculation. Similar patterns were found for PSM. Collection of SR ejaculates and dilution to 25 x 10(6)sperm/ml improved longevity of spermatozoal motility.     

Bactericidal activity of peripheral blood neutrophils during the oestrous cycle and early pregnancy in the mare

The oestrous cycles of 20 mixed-breed mares were synchronized with daily injections of 10 mg oestradiol-17 beta and 150 mg progesterone given i.m. for 10 days. On the 10th day, 10-15 mg prostaglandin F-2 alpha was administered i.m. to induce oestrus. Neutrophils were isolated from jugular blood on the 2nd or 3rd day of oestrus, Days 5 and 7 after ovulation or during early pregnancy (Days 18-34 of pregnancy). Neutrophils were challenged with Staphylococcus aureus and their bactericidal activity examined after 30 and 120 min of incubation for a reduction of colony forming units. Bactericidal activity increased with the time of incubation (P less than 0.01) but did not differ for the oestrous cycle or pregnancy (P greater than 0.05).     

Characterization of murine monoclonal antibodies directed against basic fibroblast growth factor

Monoclonal antibodies (McAbs) were developed that identify the complete (1-146 aa) and the NH2-terminal truncated (des 1-15) form of bovine basic fibroblast growth factor (bFGF). Four McAbs, designated McAbs 6, 8, 38, and 42, bind the complete form of bFGF found in bovine pituitary, brain, and adrenal gland. One of these McAbs, McAbs 42, also binds to the des 1-15 aa form of bFGF found in bovine adrenal gland, kidney, and corpus luteum. None of the McAbs binds bovine-brain-derived acidic FGF (aFGF). McAbs 6, 8, and 38 recognized the same epitope located within the first ten residues of the NH2-terminal of complete bFGF. McAb 42 recognizes a "core" epitope found on both the complete and des 1-15 aa bFGFs. The McAbs are murine IgGs with affinity constants of 10(7)-10(8) liter/M for bovine-pituitary-derived bFGF. McAbs 8 and 42 have been used in a two-site ELISA to detect the complete form of bFGF. The ELISA is sensitive to 38.5 fmole/well of bFGF and is not affected by the presence of calf serum or bovine-brain-derived aFGF. These McAbs should be useful in distinguishing the native and des 1-15 aa forms of bFGF from each other, and from aFGF and other growth factors.     

Methylcholine-activated eccrine sweat gland density and output as a function of age

The purpose of this investigation was to examine eccrine sweat gland responsiveness to intradermal injections of methylcholine (MCh) across three age groups of men [young (Y) = 22-24; middle (M) = 33-40; older (O) = 58-67 yr old, n = 5 per group]. Subjects were matched with respect to maximum O2 consumption, body size, and body composition, and were thoroughly heat acclimated before participation. Randomly ordered concentrations of acetyl-beta-methylcholine chloride ranging from 0% (saline) to 0.1% (5 x 10(-3) M) were injected into the skin of the dorsal thigh in a thermoneutral environment, and activated sweat glands were photographed at 30-s intervals for the next 8 min. Density of MCh-activated glands was independent of both age and [MCh] (e.g., 2 min after injection of 5 x 10(-3) M [MCh]: Y = 45 +/- 7, M = 46 +/- 12, O = 42 +/- 5 glands/cm2). However, sweat gland output (SGO) per active gland was significantly lower for the O group and failed to increase with increasing [MCh] above 5 x 10(-4) M. When MCh (5 x 10(-3) M) was injected after 1 h of exercise in the heat, higher SGO's were elicited in each group; however, the SGO of the O group was again significantly lower than that of the Y group (91 +/- 11 vs. 39 +/- 4 ng/gland, P less than 0.02) with the M group intermediate (69 +/- 11 nl/gland; 2 min postinjection data).(ABSTRACT TRUNCATED AT 250 WORDS)     

Evidence for an active site arginine in UDP-glucuronyltransferase

2,3-Butanedione inactivates the pure form of UDP-glucuronyltransferase used in these experiments (GT2P) (EC 2.4.1.17) purified from pig liver microsomes. The kinetics of the reaction indicates that 2,3-butanedione reacts with two amino acids that affect activity. A rapid, partial inactivation is followed by a slower rate of inactivation that leads eventually to completely inactive enzyme. UDP-glucuronic acid and glucuronic acid, as compared with UDP, are effective as protectors against the slow, secondary phase of inactivation; no ligand tested protected against the rapid phase of inactivation. The lipid environment of GT2P was a determinant of the pseudo-first order rate constant for the slow phase of inactivation, but did not affect the rate of the rapid phase of inactivation. The data suggest that GT2P contains an active site arginine that interacts with the -COO- at C-6 of the glucuronic acid moiety of UDP-glucuronic acid.     

The Epstein-Barr virus (EBV) BZLF1 immediate-early gene product differentially affects latent versus productive EBV promoters.

The Epstein-Barr virus (EBV) BZLF1 gene product is thought to mediate the disruption of latent EBV infection. We have examined the regulatory effects of BZLF1 by studying its transactivating effects on seven different EBV promoters. We find that whereas the BZLF1 gene product increases the activity of the two early promoters, BMLF1 and BMRF1, it decreases the activity of three latent promoters (the BamHI-C and BamHI-W Epstein-Barr nuclear antigen promoters and the latent membrane protein promoter). The BZLF1-induced changes in promoter-directed chloramphenicol acetyltransferase activity occur in EBV-negative as well as EBV-positive cell lines and are accompanied by a similar change in chloramphenicol acetyltransferase mRNA. Deletion analysis of the BamHI Z fragment indicates that in a portion of the amino-terminal half of the BZLF1 gene product (amino acids 24 to 86) is not essential for positive transactivating effects but is required for down-regulating effects. Thus, different domains of the same EBV immediate-early gene product can either increase the function of EBV promoters active in productive infection or decrease the function of key promoters active in latent infection.