Assay of frozen boar semen with Sephadex filtration

The suitability of filtration of frozen boar semen through Sephadex G-15-120 as a viability assay was investigated. Semen thawed on a hot plate at 38 degrees C was counted with a Coulter Counter before and after filtering through Sephadex columns with 0.1 M sodium citrate as flushing medium. More spermatozoa passed through the column when the temperature of the flushing medium was elevated from room temperature to 37 degrees C and added with 5 mM caffeine (12.3 vs. 22.8% p<0.01). The use of caffeine and 37 degrees C flushings produced a filtrate containing spermatozoa with 89+/-4% motility and 97.6+/-1.6% normal acrosome ridges with the use of frozen semen from 5 different boars. The repeatability was +/-7.3%. The unfiltered samples were judged to contain 45 +/- 8% motile spermatozoa and 66.6 +/- 7.1% spermatozoa with normal acrosome ridges. Filtering of frozen boar semen through Sephadex is proposed as a rapid, objective assay combining the benefits of differential counting of normal acrosome ridges and motility determination.     

Effect of Ca2+ and Mg2+ upon the reassociation byEscherichia coli of material released by ethylenediaminetetraacetate

Parameters involved in reconstitution of the outer membrane permeability described by Brunner, Caputo, and Treick [3] were examined. The most efficient reconstitution was obtained when divalent cations accompanied the addition of exogenous outer membrane material. Studies indicated that the effectiveness of Ca²⁺ or Mg²⁺ to promote reassociation of outer membrane material, and subsequent protection against actinomycin D, was dependent upon the strain ofEscherichia coli. More specifically, the data suggest that the effectiveness of the different divalent cations in promoting reassociation was determined by the relative amounts of F1 and F2 fractions released by ethylenediaminetetraacetate (EDTA). Reconstitution was shown by cell survival to be as high as 25% and dependent upon the total amount of material released from the outer membrane by EDTA. Between 50 and 80% of the bound material could be removed from the cells by subsequent EDTA treatment.     

Prediction of gall-stone pancreatitis by computer.

The clinical features at presentation of 53 patients admitted with primary acute pancreatitis due to gall stones were compared with those of 31 patients in whom the disease was due to other causes. Between these two groups 10 significant differences existed. By listing the frequency of symptoms and signs for each group a computer data base was prepared and incorporated into a program used in the differential diagnosis of acute abdominal pain. A program written to predict the presence of gall stones in patients with acute pancreatitis was accurate in 92% of the patients studied. A predictive index devised from the presence of three of the significantly differing clinical features correctly identified 82% of patients with gall-stone pancreatitis. Predicting the presence of gall stones on admission by analysing the presenting symptoms and signs with a computer had an accuracy comparable to that of ultrasonography or radiology and may be of value in the management of patients with acute pancreatitis.     

The effect of several diphosphonates on acid phosphohydrolases and other lysosomal enzymes.

Diphosphonates are known to inhibit bone resorption in tissue culture and in experimental animals. This effect may be due to their ability to inhibit the dissolution of hydroxyapatite crystals, but other mechanisms may be important. Since lysosomal enzymes have implicated in the process of bone resorption, we have examined the effect of several phosphonates and of a polyphosphate (P20,2) on lysosomal hydrolases derived from rat liver and rat bone. Dichloromethylene diphosphonate strongly inhibited acid beta-glycerophosphatase (EC 3.1.3.2) and acid p-nitrophenyl phosphatase (EC 3.1.3.2) and to a lesser degree (in descending order) acid pyrophosphatase (EC 3.1.3.-), arylsulfatase A (EC 3.1.6.1), deoxyribonuclease II(EC 3.1.4.6) and phosphoprotein phosphatase (EC 3.1.3.16) of rat liver. Inhibition of acid p-nitrophenyl phosphatase and arylsulfatase A was competitive. Ethane-1-hydroxy-1, 1-diphosphonate did not inhibit any of these enzymes, except at high concentrations. Neither dichloromethylene diphosphonate nor ethane-1-hydroxy-1, 1-diphosphonate had any effect on beta-glucuronidase (EC 3.2.1.31), arylesterase (EC 3.1.1.2) and cathepsin D (EC 3.4.23.5). Of several other phosphonates tested only undec-10-ene-1-hydroxy-1, 1-diphosphonic acid inhibited acid p-nitrophenyl phosphatase strongly, the polyphosphate (P20, I) had little effect. Acid p-nitrophenyl phosphatase in rat calvaria extract behaved in the same way as the liver enzyme and was also strongly inhibited by dichloromethylene diphosphonate, but not by ethane-1-hydroxy-1, 1-diphosphonate. It is suggested that the inhibition of bone resorption by dichloromethylene diphosphonate might be due in part to a direct effect of this diphosphonate on lysosomal hydrolases.     

The location of proteins labeled by the 125I-lactoperoxidase system in the NIL 8 hamster fibroblast.

NIL 8 hamster fibroblast cells were labeled by lactoperoxidase-catalyzed iodination. Their membranes were fractionated by sedimentation-rate and isopycnic zonal centrifugation. All the iodinated proteins except the very prominently labeled high molecular weight protein (greater than 200,000 daltons) were located in a fraction identified enzymically and compositionally as plasma membrane. The high molecular weight protein that was previously shown to be sensitive to virus transformation (Hynes, 1973) is concentrated in a very high density particle (rho equals 1.253-1.259) which contains mainly carbohydrate and protein and very low levels of lipid. 5'-nucleotidase was the only enzyme reproducibly demonstrated in this fraction, and electron micrographs revealed a predominantly amorphous morphology together with a few membraneous structures. The iodine label in this fraction was very sensitive to trypsinization prior to homogenization. All the available evidence indicates that this fraction is derived from the surface coat. Mitochondria, nuclei, and soluble protein were labeled to an insignificant extent. The presence of the iodinated surface proteins associated with the endoplasmic reticulum fraction is discussed in the light of these results.     

Clearance of bacteria and non-antigenic markers following intra-uterine inoculation into maiden mares: Effect of steroid hormone environment

Uterine response to inoculation with Streptococcus zooepidemicus organisms (antigenic markers) and 15-mum microspheres and charcoal (non - antigenic markers) was determined in seasonally acyclic maiden mares treated with either progesterone (P) n = 4, estradiol (E) n = 4 or oil vehicle (C) n = 4. At 3, 7 and 15 d after inoculation with bacteria and the 2 non - antigenic markers, uteri were flushed and the clearance of these materials, as well as the number of white blood cells and immunoglobulin concentration, determined. P-treated mares had higher numbers of bacteria and IgA and a greater volume of purulent fluid in the uterus than E- or C-treated mares at 7 d after inoculation. Clearance of inoculated materials began within 2 h in E-treated mares, and the non-antigenic markers were completely cleared in E- and C-, but not in P-treated mares, in 3 d. This suggests that in the P-dominated uterus, reduced physical clearance may contribute to an increased susceptibility to uterine infection.     

Is There a Genetic Basis for the Deposition of β-Amyloid After Fatal Head Injury?

1. Alzheimer's disease is a heterogeneous disorder that may be caused by genetic or environmental factors or by a combination of both. Abnormalities in chromosomes 1, 14, and 21 have all been implicated in the pathogenesis of the early-onset form of the disease, while the 4 allele of the apolipoprotein E gene (on chromosome 19) is now recognized as a risk factor for early- and late-onset sporadic and familial Alzheimer's disease.2. The best-established environmental trigger for the disease is a head injury, based on epidemiological and neuropathological evidence. Approximately 30% of patients who die after a single episode of severe head injury show intracerebral deposition of -amyloid protein (A), a protein that is thought to be central to the pathogenesis of Alzheimer's disease.3. Recent studies have revealed an over-representation of the apoE 4 allele in those head-injured patients displaying A pathology, thus providing the first evidence for a link between a genetic susceptibility (apoE 4) and an environmental trigger (head injury) in the development of Alzheimer-type pathology.