Comparison of pig and chicken pepsins for protein hydrolysis.

The aim of the present study was to compare the degree of proteolysis with pig (PP) and chicken (CP) pepsins in order to find out whether PP can be used instead of CP to simulate gastric hydrolysis in the chicken. First, the pH activity profile of the two pepsins was compared using three substrates. For haemoglobin, CP showed a slightly higher optimal pH than PP, 2.5-3 and 2, respectively. For two plant protein sources (peas, wheat), the optimal pH was similar for the two enzymes, about pH 1.5. For the three substrates tested, CP exhibited a high level of activity over a broader pH range than PP. Second, the susceptibility of the two plant proteins to hydrolysis by each of the two pepsins was studied at pH levels near the chicken gastric pH (1.5-3.5). For PP, pea proteins were hydrolysed more than wheat ones, while, for CP, the hydrolysis was dependent on pH. Therefore, the classification of the two studied protein sources was dependent on the enzyme species and pH. The results of this study show that the choice of in vitro hydrolysis conditions to assess the digestibility of proteins must be made with great care.

Increased insulin receptor number and insulin responsiveness in a chicken hepatoma cell line.

Insulin receptor number and insulin responsiveness were compared in a chicken hepatoma cell line (LMH) and in normal chicken hepatocyte (cHep) cells cultured in the same conditions. LMH cells expressed two- to threefold more insulin receptors than cHep cells, without significant changes in affinity. The tyrosine kinase activity of solubilized and lectin (lentil+wheat germ agglutinin; WGA)-purified LMH receptors was higher than that of cHep receptors. The ATP hydrolytic activity previously observed in WGA-purified receptors from chicken liver membranes was also present in WGA-purified receptors from cultured cHep cells. This unidentified membrane-associated ATPase was absent from LMH membrane-solubilized material and therefore from WGA-purified LMH insulin receptors. Finally, LMH cells incorporated at least tenfold more amino isobutyric acid than cHep cells in the absence of insulin and were more responsive to insulin. The enhanced basal amino acid transport of LMH cells was most probably the consequence of their proliferative activity. The enhanced insulin responsiveness of LMH cells can be accounted for, at least in part, by one or several of the modifications presently demonstrated in LMH cells when compared with normal cultured hepatocytes: increased insulin receptor number and tyrosine kinase activity and possibly the loss of the membrane-associated ATPase.

Insulin receptor and insulin sensitivity in a chicken hepatoma cell line.

Insulin receptors have been characterized in a cell line recently isolated from a chicken hepatoma (LMH). The binding of 125I-insulin to LMH cells or membranes displayed the expected criteria for insulin receptors: affinity, temperature dependency, curvilinearity of Scatchard plot, rank order of potency for insulin analogs and insulin induced down-regulation. The alpha-subunit of LMH cell insulin receptors exhibited a normal size of 135 kDa. Following autophosphorylation, LMH WGA-purified receptors revealed a 95 kDa beta-subunit and a 72 kDa protein (pp72). Both proteins were phosphorylated in a time-, insulin- (and insulin-like growth factor 1; IGF-1) and manganese-dependent manner, and were precipitated by antiphosphotyrosine and two anti-insulin receptor antibodies. The 72 kDa protein was not present under non-reducing condition PAGE or in normal chicken liver. These results strongly suggest that pp72 is either a truncated form of the insulin receptor beta-subunit specific to LMH cells or a degradation product. Lectin-purified insulin receptors from LMH cells or chicken liver membranes exhibited similar tyrosine kinase activity, using artificial substrate poly(Glu-Tyr) 4:1. Finally, amino acid uptake by LMH cells was insulin stimulatable.

'Serum like' albumin of fowl seminal plasma and effects of albumin on fowl spermatozoa stored at 4 degrees C.

1. Using immunoelectrophoretic and immunodiffusion methods, serum-like albumin was detected in fowl seminal plasma. Immunodiffusion showed seminal plasma albumin concentration to be 4 mg/ml, corresponding to half of the total proteins (8 mg/ml). 2. Replacing seminal plasma with a diluent containing either 1, 4, or 16 mg/ml albumin increased motility of spermatozoa stored for 24 h at 4 degrees C, 16 mg/ml being the more effective dose. 1 and 4 mg/ml had no effect on the fertilising ability of fowl spermatozoa stored for 24 h at 4 degrees C in both young (28-35 weeks old) and old birds (50-55 weeks old). 16 mg/ml albumin had no effect on fertilising rates in young but depressed it in old birds. 3. These results indicate that seminal plasma albumin may be one of the mobility stimulating factors of seminal plasma. However it does not protect fertilising ability better than the diluent alone.

[Polyradiculoneuritis following lumbosacral zona with adrenal cortex adenoma]. / Polyradiculonévrite secondaire à un zona lombo-sacré avec adénome cortico-surrénalien.

We report a case of severe polyradiculoneuritis consecutive to a lumbosacral herpes zoster and concomitant with an adrenal adenoma with hypercorticism. The patient improved after surgery and plasmaphereses.

Physiological and pathological factors influencing bovine immunoglobulin G2 concentration in milk.

Bovine IgG2 concentration was determined by radial immunodiffusion in 355 milk samples of uninfected quarters, 101 milk samples of infected quarters, and 118 blood serum samples from 42 Holstein-Friesian cows taken at 30, 150, and 270 d. Concentration of IgG2 in blood serum (11.3 mg/ml) was highest at the beginning of lactation (30 d). Immunoglobulin G2 concentration in milk (16.81 micrograms/ml) from cows with uninfected quarters was not affected by quarter location but was correlated with IgG2 concentration in blood serum (.30; P less than .001). The IgG2 concentration in milk was lower in midlactation (150 d: 14.81 micrograms/ml) and in the two first lactations. Immunoglobulin G2 concentration in milk was correlated with SCC. Quarter infection by Corynebacterium bovis or major pathogens increased IgG2 concentration up to 47.9 micrograms/ml for Staphylococcus aureus. Only S. aureus influenced IgG2 concentration in blood serum. Correlation between IgG2 content and SCC in milk decreased when quarters were infected, regardless of bacterial species.

Role of milk immunoglobulins in the Brucella milk ring test.

Milk immunoglobulins were extracted from the stained cream layer of positive milk ring tests from experimentally inoculated or naturally infected cows. IgA was always found, associated with IgM in most cases (15/17) and with IgG in a smaller number of cases (11/17). An additional incubation at 20 degrees C for 18 h gave clearer positive and negative results and a lower limit of detection than that of the usual milk ring test.

Physiological and pathological factors influencing bovine alpha-lactalbumin and beta-lactoglobulin concentrations in milk.

Bovine alpha-lactalbumin and beta-lactoglobulin concentrations were determined by radial immunodiffusion in 354 milk samples from uninfected and 98 samples from infected quarters from 42 Holstein-Friesian cows taken at 30, 150, and 270 days of lactation. alpha-Lactalbumin and beta-lactoglobulin concentrations were not affected by quarter location. The alpha-lactalbumin decreased at the end of lactation and in samples collected beyond second lactation. The beta-lactoglobulin concentration increased with stage of lactation. There was a positive correlation between alpha-lactalbumin and beta lactoglobulin (r = .12). Milk from uninfected quarters had mean alpha-lactalbumin and beta-lactoglobulin concentrations of 1.47 and 4.6 mg/ml, respectively. Milk from quarters infected by major pathogens or Corynebacterium bovis had less alpha-lactalbumin. Milk from quarters infected by minor pathogens had less beta-lactoglobulin. There was a negative correlation between alpha-lactalbumin concentration and somatic cell count (r = .31), which was amplified by infection status of quarters. No correlation was noted between somatic cell count and beta-lactoglobulin concentration when considered over the whole sampling period, but the correlation became negative in quarters infected by major pathogens.

Assessment of hemolytic and bactericidal complement activities in normal and mastitic bovine milk.

Bactericidal and hemolytic complement activities were investigated in 51 quarter milk samples of 13 cows in late lactation. Hemolytic activity was in all of the samples but one, after accounting for whey inhibitory activity. Mean hemolytic activity and inhibitory activities were .18 and .34 complement hemolytic units. Inflammation, in relation to infection status, increased hemolytic titers and heat-labile bactericidal activities of milk. Correlation coefficient was .76 between albumin content of milk serum and hemolytic titer of samples from infected quarters. Normal milk decreased bactericidal titer of bovine serum against a serum-sensitive Escherichia coli strain. When compared to veronal buffer saline solution, milk did not accelerate decay of hemolytic activity over 7-h incubation at 39 degrees C. Taken together, these results suggest that the adverse effect of milk on both hemolytic and bactericidal activities of complement is limited and might be of significance essentially before full establishment of the inflammatory reaction to bacterial invaders.

A sensitive microassay for the determination of hemolytic complement activity in bovine milk.

A 51Cr release microhemolytic complement assay is described to detect hemolytic complement activity in bovine milk. 51Cr-labeled guinea-pig erythrocytes (GPRBC), which have been sensitized with a subagglutinating amount of rabbit anti-GPRBC, are placed in microtiter plates. Pooled bovine sera as source of complement to achieve about 50% of 51Cr release were added to each well prior to the addition of the samples on the test. Determination of CH100 titer was obtained by difference of counting between heated and unheated diluted whey samples from a standard linear regression. Comparative hemolytic values throughout lactation were established for the first time and confirmed the improved sensitivity of the assay.

Physiological and pathological factors influencing bovine immunoglobulin G1 concentration in milk.

Bovine immunoglobulin G1 concentration was determined by radial immunodiffusion in 349 milk samples of uninfected quarters, 95 of infected quarters, and 118 blood serum samples from 42 Holstein-Friesian cows taken at days 30, 150, and 270. In lactation, immunoglobulin G1 concentration in milk was not affected by immunoglobulin G1 concentration in blood serum or location of quarters. The immunoglobulin G1 concentrations increased at the end of lactation and in samples collected from cows beyond the third lactation. Uninfected quarters had a mean immunoglobulin G1 concentration of .46 mg/ml. This was less than means from quarters infected by minor or major pathogens. Quarter infection by Staphylococcus aureus resulted in an increase of immunoglobulin G1 concentration in blood serum (9.22 to 11.3 mg/ml). When Corynebacterium bovis was persistent throughout the lactation, immunoglobulin G1 concentration in blood serum was increased (11.26 mg/ml). There was no correlation between somatic cell count and immunoglobulin G1 concentration in uninfected quarters. There was a slight correlation between bovine serum albumin and immunoglobulin G1 concentration in identical quarters (.23). Infection of quarters increased in varying degrees the correlation between immunoglobulin G1 concentration and bovine serum albumin concentration or somatic cell count in milk.

Physiological and pathological factors influencing bovine serum albumin content of milk.

Concentrations of albumin from bovine blood serum were measured by radial immunodiffusion of 480 milk samples. Content in milk was not affected by content in serum, lactation number, or location of quarters. Bovine serum albumin concentrations in milk increased at the end of lactation (270 days) compared to the beginning (30 days) and at midlactation (150 days). Uninfected quarters had a mean concentration of serum albumin of .193 mg/ml. This was less than means for quarters infected by minor pathogens (.242 mg/ml) and by major pathogens (.284 mg/ml). Distributions of concentrations related to infection status, however, overlapped substantially. Somatic cell count was correlated .53 with concentrations of blood serum albumin in milk. About 32% of quarters infected by major pathogens had fewer than 500 x 10(3) cells/ml, whereas 47.5% of them had serum albumin content less than .2 mg/ml. For subclinical infections, concentration of serum albumin markedly increased when somatic cells were more than 1,000 x 10(3) cells/ml.

Sequential changes in serum albumin, immunoglobulin (IgG1, IgG2, IgM) and lactoferrin concentrations in milk following infusion of Escherichia coli into the udder of immunised and unimmunised cows.

Two immunised and three unimmunised cows were infected in a single mammary gland with 10(4) CFU of the vaccine Escherichia coli strain. Immunisation comprised systemic (subcutaneous) injection of killed bacteria at drying-off and one intramammary infusion five weeks later. Immunoglobulin (Ig) G1, IgG2, lgM, serum albumin (BSA) and lactoferrin concentrations were monitored by sampling the inoculated glands at 2 h-intervals during the first 16 h post-inoculation, then at each milking for four days. Whether immunised or not, mammary glands started to react at 10 h post-inoculation. During the early acute phase, IgG1 and IgG2 permeated from blood into milk at a rate similar to BSA. Later on, IgM (and at a lower degree IgG1) concentrations were higher than expected on the basis of passive transfer. Marked protein exudation was seen in all of the cows but one. Nevertheless, this cow (immunised) showed an intense cellular reaction like the other animals. Lactoferrin concentrations rose from 24-32 h post-inoculation and remained elevated to the end of the observed period in inoculated quarters in unimmunised cows. By contrast, in immunised cows lactoferrin concentrations remained low. Heat-labile bactericidal activity against a serum-sensitive E. coli strain appeared concomitantly with rise in BSA concentration. Heat-resistant bactericidal activity of cell-free milk was detected one or two days later in three of the cows. Bacteriological cure of quarters occurred without therapy in all cases.

Lactoferrin and transferrin in bovine milk in relation to certain physiological and pathological factors.

Mean transferrin and lactoferrin concentrations in whey samples from 376 uninfected quarters of 42 Holstein X Friesian cows on the 30th, 150th and 270th days of lactation were respectively 0.030 and 0.080 mg/ml. The mean transferrin concentration in serum was 4.63 mg/ml. Lactation number and location of quarters did not influence milk lactoferrin and transferrin values. Lactoferrin concentration increased significantly (P less than 0.01) in uninfected quarters from 0.03 to 0.06 and to 0.15 mg/ml on the three successive sampling times. Transferrin whey concentration increased significantly (P less than 0.01) only in late lactation, from 0.025 (30th and 150th days of lactation) to 0.035 mg/ml (270th day of lactation). Lactoferrin concentration increased significantly (P less than 0.01) in quarters infected by major pathogens (41 samples) whereas minor pathogen infections (61 samples) caused no significant increase. The correlation coefficients between milk lactoferrin and transferrin concentrations and somatic cell-count were 0.42 and 0.65 respectively.

Lysostaphin disk test for routine presumptive identification of staphylococci.

The sensitivity of the lysostaphin disk test was evaluated for routine differentiation of staphylococci from micrococci. Lyophilized paper disks impregnated with 10 micrograms of lysostaphin were placed on a Mueller-Hinton agar plate inoculated with 10 species of staphylococci and 7 species of micrococci. After 16 h of incubation at 37 degrees C, a circular zone of growth inhibition was noted for all staphylococci species tested. By contrast, all the micrococci species were resistant. The activity of the impregnated disks was not diminished by storing them at 4 degrees C for 3 months. With disks impregnated by 10, 6.6, 5, and 2.5 micrograms of lysostaphin, diameters of the growth inhibition zone for S. aureus Cowan 1 NCTC 8530 strain were proportional to the logarithm of the quantity of lysostaphin present in each disk.

Les hemoglobines des amphibiens: Separation et caracterisation preliminaire des chaines d'une hemoglobine du crapaud bufo bufo.

A toad (Bufo bufo) hemoglobin has been purified and two types of chain have been separated by counter-current distribution. The alpha-chain has a N-terminal sequence Ac-Ala-Leu and a C-terminal sequence Tyr-Arg. The beta-chain has a N-terminal sequence Val-His-Leu and a C-terminal sequence Tyr-His.