CD34 glycoprotein identifies putative stem cells located in the isthmic region of canine hair follicles

It is widely documented that a pool of multipotent stem cells located in humans and mice hair follicle outer root sheath (bulge region) is involved in the restoration of the whole follicular unit during each anagen phase. To the authors' knowledge, data regarding the location and characterization of hair follicle stem compartment in dogs have not been reported in the recent relevant literature. In this study, we investigated the haematopoietic stem and progenitor cell antigen CD34 as a marker of putative stem cells located in a bulge-like region of canine hair follicles. The presence of CD34 mRNA and glycoprotein was assessed on formalin-fixed, paraffin-embedded canine skin samples by in situ hybridization technique and by standard immunohistochemistry, respectively. A strong expression of CD34 mRNA and glycoprotein was observed in a well-defined area of the hair follicle isthmic region and appeared uniformly concentrated at the level of the basal layer of the outer root sheath. These findings provide compelling support to the hypothesis that in dogs, a subpopulation of basal keratinocytes located in the hair follicle isthmic region and characterized by the selective expression of CD34 is potentially associated with the stem cell compartment of this skin appendage.     

Evaluation of young and adult dogs' reactivity

Many authors have tested puppies' behavior early to detect adult behavioral disorders. Our pilot study aims to investigate puppies' reactivity toward tolerance of new situations and the consistency of behavior between puppy and adult dog. Seven tests (tunnel, direct looking, sudden noise, retrieving buster-ball, problem solving, bowl removal, and approaching) were used to evaluate the reactivity of 32 German shepherd dogs at 5, 7, and 9 months of age. Among these dogs, 17 were chosen by physical and behavioral traits to continue the training program up to adulthood (choice group) and administered the same tests at the age of 2 years. Through the observation of behaviors, the tests aim to assess sociality, exploratory and retrieving tendency, learning ability, playing aptitude, and tendency toward offensive aggression. The behaviors were ranked by degree of reactivity, and ranks were used as scores. The effects of gender and group on the 5- to 9-month reactivity profiles of all the puppies were estimated with a repeated-measures model. In the choice group, the behavior changes from 9 to 24 months and their dependence on gender were estimated with an analogous model. All puppies became more self-confident between 5 and 9 months. At 9 months, behavior was slightly better in males and in the choice group. Between 9 and 24 months, self-confidence, attentiveness, and ability to solve problems and retrieve objects improved, whereas playfulness decrease. Behaviors at 9 and 24 months showed very low variability, which indicated their very high similarity.     

Histological and immunohistochemical detection of different Helicobacter species in the gastric mucosa of cats.

Detailed histopathological evaluation of the gastric mucosa of Helicobacter-infected cats is complicated by the difficulty of recognizing Helicobacter organisms on hematoxylin and eosin (HE)-stained sections and the ability of multiple Helicobacter species to infect cats. In this study, the presence and localization of different species of Helicobacter in the stomachs of cats was investigated using silver staining and immunohistochemistry. Five groups containing 5 cats each were established (group 1: urease negative and Helicobacter free; groups 2, 3, 4, and 5: urease positive and infected with Helicobacter heilmannii, unclassified Helicobacter spp., Helicobacter felis, and Helicobacter pylori, respectively). Gastric samples were evaluated by HE and silver staining and by immunohistochemistry with 3 different anti-Helicobacter primary antibodies. Helicobacter were detected by Steiner stain in all infected cats at the mucosal surface, in the lumen of gastric glands, and in the cytoplasm of parietal cells. In silver-stained sections, H. pylori was easily differentiated from H. felis, H. heilmannii, and unclassified Helicobacter spp., which were larger and more tightly coiled. No organisms were seen in uninfected cats. Helicobacter antigen paralleled the distribution of organisms observed in Steiner-stained sections for 2 of the 3 primary antibodies tested. The antisera were not able to discriminate between the different Helicobacter species examined. A small amount of Helicobacter antigen was present in the lamina propria of 3 H. pylori-, 3 H. felis-, and 1 H. heilmannii-infected cat. Minimal mononuclear inflammation was present in uninfected cats and in those infected with unclassified Helicobacter spp. and H. heilmannii cats. In H. felis-infected cats, lymphoid follicular hyperplasia with mild pangastric mononuclear inflammation and eosinophilic infiltrates were present. The H. pylori-infected cats had severe lymphoid follicular hyperplasia and mild to moderate mononuclear inflammation accompanied by the presence of neutrophils and eosinophils. These findings indicate that Steiner staining and immunohistochemistry are useful for detecting Helicobacter infections, particularly when different Helicobacter species can be present. Monoclonal antibodies specific for the different Helicobacter species could be important diagnostic aids. There appear to be differences in the severity of gastritis in cats infected with different Helicobacter species.     

Fermentability of grape must after inhibition with dimethyl dicarbonate (DMDC)

Dimethyl dicarbonate (DMDC) was added to grape must and to synthetic media and results showed that, at 20 degrees C, 150 mg.L(-)(1) DMDC completely inhibited the fermentation of a grape must that was previously inoculated with 10(6) cells.mL(-)(1) Saccharomyces bayanus and Saccharomyces uvarum. Brettanomyces intermedius, Candida guilliermondii, Hansenula jadinii, Hansenula petersonii, Kloeckera apiculata, Pichia membranaefaciens, and Saccharomyces cerevisiae were inhibited by 250 mg.L(-)(1). Candida valida was inhibited in the presence of 350 mg.L(-)(1), whereas Hanseniaspora osmophila, Saccharomycodes ludwigii, Schizosaccharomyces pombe, and Zygosaccharomyces bailii required 400 mg.L(-)(1). Delay of fermentation (but not inhibition) was noted in the presence of 400 mg.L(-)(1) for the following cultures: Brettanomyces anomalus, Hanseniaspora uvarum, Metschnikowia pulcherrima, Schizosaccharomyces japonicus, Torulaspora delbrueckii, and Zygosaccharomyces florentinus. Acetobacter aceti and Lactobacillus sp. were completely inhibited using 1000 and 500 mg.L(-)(1) DMDC, respectively. The fermentation of a grape must inoculated with 10(6) cells.mL(-)(1) of different wine yeasts was delayed for 4 days after the prior addition of 200 mg.L(-)(1) of DMDC; 200 mg.L(-)(1) DMDC did not show any residual inhibitory effect after 12 h, nor did 300 mg.L(-)(1) 24 h after the addition. In cellar experiments, indigenously contaminated grape musts (with and without skins) showed a delay in fermentation of 48 h after the addition of only 50 mg.L(-)(1) DMDC. The possibility of using DMDC (as pure grade as commercially available) in grape must as a disinfectant for the decontamination of musts indigenously contaminated with wild yeast should be considered seriously, despite its apparent low solubility in water.     

Characterization of "lettucine", a serine-like protease from Lactuca sativa leaves, as a novel enzyme for milk clotting

In this work we focused on the characterization of a novel plant rennet purified from lettuce leaves (Lactuca sativa L. cv Romana). The lettuce protease, lettucine, showed trypsin-like, SV8-like, and caseinolytic activities. Although the enzyme did not recognize peptides having hydrophobic amino acid residues in the P(1) position of the target bond, it did show milk-clotting activity, suggesting that different bonds rather than the Phe(105)-Met(106) of the kappa-casein might be cleaved, still inducing milk-clotting. The enzyme exhibited proteolytic activity toward alpha-casein, beta-casein, kappa-casein, and milks with different fat contents, with the highest activity observed with partially skimmed milk, total casein, and alpha- and kappa-casein. SDS-PAGE studies showed that lettucine cleaved alpha-casein, beta-casein, and kappa-casein. In particular, we showed that alpha-casein breakdown occurred even though total casein or milks were supplied, suggesting that the lettuce enzyme is able to operate a significant disorganization of the casein's micellar structure. Moreover, the proteolytic activity of the enzyme analyzed under various technological parameters, such as temperature and pH, indicated that the lettuce enzyme is highly consistent with the milk-clotting process.     

Comparison of gel column agglutination with monoclonal antibodies and card agglutination methods for assessing the feline AB group system and a frequency study of feline blood types in northern Italy

Background: A new commercial gel column agglutination system is reported to have high sensitivity in detecting cats with blood type AB. Objectives: The aims of this study were to compare gel column agglutination and card agglutination methods for feline blood‐typing and to determine the frequency distribution of feline blood types in northern Italy. Methods: Blood‐typing was performed on 120 cats using both a commercial gel column containing monoclonal antibodies (ID Gel‐Test Micro Typing System) and a card agglutination method (RapidVet‐H Feline). Results were confirmed with back‐typing. Sensitivity, specificity, positive predictive value, and negative predictive value were calculated for the 2 methods. A second group of 140 Domestic Shorthair (DSH) cats was blood‐typed using the gel column technique to determine the frequency distribution of feline blood types in northern Italy. Results: The card agglutination method demonstrated poor sensitivity in identification of type‐AB cats (61%) and was only 95% specific when identifying type‐B cats. The gel column agglutination technique demonstrated 100% sensitivity and specificity for typing all 3 blood types (A, B, and AB). The frequency distribution study of 140 cats demonstrated that 127 (90.7%) cats were type A, 10 (7.1%) were type B, and 3 (2.1%) were type AB. Conclusion: When blood‐typing cats of breeds with a relatively high frequency of blood types B and AB, methods that use monoclonal antibodies for detection of blood types B and AB are recommended. Alternatively, blood type can be confirmed by more sensitive supplemental testing, such as back‐typing. The high frequency of blood type A in DSH cats in northern Italy was comparable to previously reported frequencies in Italy and world‐wide.     

Physical, Hematological, and Biochemical Responses to Acute Intense Exercise in Polo Horses

The aim of this study was to evaluate the response of physical, hematological, and biochemical parameters after acute intense exercise in polo horses playing in an outdoor international competition. The game consisted of four periods (chukkas) and each period consisted a playing time of 7 minutes. Two matches were played everyday for a week. A total of 12 horses were examined. Each “high-goal” polo horse played one chukka a day for 4 days. Horses were clinically examined the day before the games started and then daily during the 4 days of their participation in the games. During these days, physical examination was performed and blood sample was collected at rest (T0), immediately (T1) after exercise, and after 30 minutes of exercise (T2). Blood samples were analyzed for total cell counts and for determination of creatine kinase, lactate dehydrogenase (LDH), aspartate aminotransferase, lactate, total proteins, calcium, magnesium, phosphorus , and cortisol. Data were evaluated using two-way analysis of variance. Exercise caused significant dehydration (P < .01), mucous membranes congestion, increased heart rate (P < .001), and capillary refill time (P < .001). It also caused increased value of the following parameters: hematocrit (P < .001), red blood cells (P < .001), hemoglobin (P < .001), white blood cells (P < .05), lymphocyte (P < .001), total proteins (P < .001), creatine kinase (P < .05), LDH (P < .01), lactate (P < .001), and cortisol (P < .01), and a decrease in the platelet count (P < .001), calcium (P < .01), phosphorus (P < .001), and magnesium (P < .001). All parameters returned within or near the reference range by 30 minutes postexercise. On the basis of these observations, data were considered indicative of a good response to an acute intense exercise. Moreover statistical results obtained were typical of a mixed aerobic/anaerobic metabolic pathway that is prevailing in this sport.