Stefania Casella

Production of canine mesenchymal stem cells from adipose tissue and their application in dogs with chronic osteoarthritis of the humeroradial joints.

Autologous AD‐MSC [adipose‐derived MSC (mesenchymal stem cell)] therapy involves harvesting fat from the patient by isolating the stem and regenerative cells and administering the cells back to the patient. This study evaluated the production of canine AD‐MSCs and their possible application in cellular therapy for dogs. To assess whether cellular therapy can replace drug therapy, the clinical effect of a single intra‐articular injection of AD‐MSCs was evaluated on 4 dogs with lameness associated with OA (osteoarthritis) of the humeroradial joints. MSCs were readily isolated from adult dog adipose tissue, and their ability to form colony and differentiate into various phenotypes was confirmed. AD‐MSCs expressed OCT4, NANOG and SOX2 at the mRNA level, pluripotency markers usually ascribed to embryonic stem cells. The results suggest the stemness of the cells isolated from canine fat, and good quality control made them available for both experimental and clinical use. Follow‐up studies to evaluate the effects of AD‐MSC therapy showed that OA of the elbow joints improved with time, indicating significant potential for clinical use in the treatment of lameness, particularly when administered before the injury becomes severe.

Analysis of serum proteins in clinically healthy goats (Capra hircus) using agarose gel electrophoresis

BACKGROUND Electrophoretic patterns of serum proteins provide useful information on pathological conditions in ruminants. Their reference values, however, are dissimilar to those of other species. Reference values for goats using agarose gel as the supporting matrix have not been reported. OBJECTIVE The aim of this study was to evaluate serum concentrations of total protein and protein fractions (albumin and globulins) by means of agarose gel electrophoresis (AGE) in goats in order to establish electrophoretic reference intervals and to evaluate potential changes associated with aging. METHODS Blood was collected from 105 clinically healthy Girgentana goats by means of jugular venipuncture. Serum protein concentrations were assessed by AGE. Three age groups were compared: 1-1.5 years, 2-4 years, and 5-12 years. RESULTS Values (mean ± SD) were determined for concentrations of total protein (72.26 ± 6.40 g/L), albumin (31.80 ± 4.00 g/L), α-globulins (6.40 ± 1.23 g/L), β(1) -globulins (10.50 ± 2.58 g/L), β(2) -globulins (5.18 ± 1.60 g/L), and γ-globulins (18.65 ± 5.90 g/L) and for albumin/globulin (A/G) ratio (0.82 ± 0.20). One-way ANOVA showed statistically significant age-related differences for total protein and α-globulin concentrations and A/G ratios. Age influenced protein concentrations with the 5-12-year-old group having higher total protein and α-globulin concentrations and lower albumin concentration and A/G ratios than the 2-4-year-old group. CONCLUSIONS This study provides reference values for total protein concentrations and protein fractions obtained by AGE in goats. Some values vary with age. Age-specific reference intervals are reported in order to provide clinicians with an additional diagnostic aid.

Canine mesenchymal stem cells (MSCs): characterization in relation to donor age and adipose tissue-harvesting site.

Adipose tissue as a stem cell source is ubiquitously available and has several advantages compared to other sources, for example it is easily accessible in large quantities with minimal invasive harvesting procedure, and isolation of adipose‐derived mesenchymal stem cells (MSCs) yields a high amount of stem cells, essential for stem cell‐based therapies and tissue engineering. We have explored the effect of donor age, and the anatomical origin of the adipose tissue on several aspects of MSCs in dogs, such as cell yield, proliferative ability, multi‐differentiation potential, colony‐forming capacity, stemness marker expression. We also assessed the effect of cell passaging on the MSCs stemness. We found that the anatomical origin of the adipose tissue and the age of donors have effects only on the proliferative capacity of the MSCs. Moreover, cells show a progressive loss of the stemness characteristics with passages. Cell therapies need a suitable number of cells to use in clinical applications. Characterization of MSCs at different passages, allowed us to demonstrate that, under our culture conditions, the best quantitative and qualitative characteristics are obtained at early passages. Adult MSCs are of particular interest for the therapeutic approach to musculoskeletal diseases, and the dog provides an excellent preclinical model for the development of new approaches in regenerative medicine that might be applied to humans.

Influence of transportation on serum concentrations of acute phase proteins in horse

The modifications of Haptoglobin (Hp), Serum Amyloid A (SAA), Fibrinogen (Fbg) and White Blood Cells (WBCs) were evaluated in 15 Saddle Italian horses. Ten horses were transported covering a distance of about 320 km within 4 h with an average speed of 80 km/h (experimental group) and five horses were not subject to transportation (control group). Blood was collected via jugular venipuncture before the transportation (T0), immediately after the transportation (T1), 12 (T12), 24 (T24) and 48 (T48)hours after the transportation in experimental group and at the same time point in control group. For each parameter statistical analysis of different groups and sampling time was performed using a two-way analysis of covariance, with the data before the transportation (T0) as the covariate, by the GLM procedure of SAS. For all parameters the interaction (Group × Time) was tested and it was resulted no significant. The application of statistical analysis showed significant differences between the control group and horses subjected to transportation (P<0.01), and the influence of sampling time (P<0.05) on Hp, SAA and WBCs. These modifications appeared to be innovative showing that equine Hp, generally considered as moderate acute phase protein, increases more rapidly than the SAA after transportation-induced stress.

Modifications of some acute phase proteins and the white blood cell count in thoroughbreds during training

The concentrations of plasma fibrinogen and serum haptoglobin and the white blood cell count (WBC) were assessed in 17 thoroughbreds. All horses were trained for 80 days, six days a week, with one day of rest per week. Blood samples were collected from each animal by jugular venepuncture, at 06.00, every 20 days for a period of 80 days, and the fibrinogen and haptoglobin concentrations and WBC were determined. One-way repeated measures analysis of variance, followed by the Bonferroni test, was used to determine significant differences. Training was associated with a significant increase in the concentrations of plasma fibrinogen (Plt;0.01) and serum haptoglobin (Plt;0.0005), but had no significant effect on the WBC count.

Influence of age on profile of serum proteins in the calf.

The authors studied the course of serum proteins during the first week and the first month of life in the calf in order to obtain useful information for neonatal care. Eight Limousine calves, four males and four females, clinically healthy, were used. From all animals blood samples were collected from the external jugular vein from the day after the birth every day for one week and every five days for thirty days. Blood samples were taken at the same hour (09:00), and the serum concentrations of albumin, globulins (α1-globulins, α2-globulins, β- globulins, γ-globulins) and total proteins were determined by means of automated multiparametric agarose gel electrophoresis system. One-way repeated measures analysis of variance (ANOVA), followed by the Bonferronis test, was used to determine significant differences. Data analysis of variance showed a significant effect of days of life, with p < 0.05, only on albumin, α1-globulins and α2-globulins during the first month of life. These results confirm that despite the fact that newborns must adapt to various environmental factors after birth, including nutrition that can influence the serum protein profile during neonatal period, the evaluation of electrophoretics parameters during the first months of life results useful for the diagnosis and treatment of neonatal diseases.

Annual rhythms of some physiological parameters in Ovis aries and Capra hircus

The aim of this study was to evaluate the annual variation of some physiological parameters (total lipid, NEFA, phospholipids, triglycerides, creatinine, β-idrossibutirrate, albumin, total protein, total cholesterol, urea, uric acid, glucose, rectal temperature and heart rate) in five female goats and five female sheep housed under natural photoperiod and natural ambient temperature in a 12 m2 sound proof box equipped with 50 × 100 cm opening window, that allowed natural ventilation. ANOVA was used to determine significant differences (p values < 0.05 were considered statistically significant). A trigonometric statistical model was applied to describe the periodic phenomenon analytically, by individuating the main rhythmic parameters according to the single cosinor procedure. Serum total lipid, NEFA, phospholipids, triglycerides, rectal temperature and heart rate, both in sheep and goats, showed an annual rhythm with acrophases in autumn. The results show that sheep and goats present similar annual patterns, probably influenced by various environmental factors.

Pattern of serum protein fractions in dairy cows during different stages of gestation and lactation

In dairy cows the period of transition from late gestation to early lactation is recognized as inducing considerable metabolic adaptation. The aim of this study was to analyse modifications in serum protein values occurring during the dry and the transition period and during lactation in a group of five Holstein cows of high average milk production. For all subjects, selected on the basis of their pregnancy status, blood samples were collected at different physiological phases: dry period (-60, -30 d to calving), transition period (almost 7 d to calving, 7 d after calving), and lactation (weeks 2, 5 and 15 after calving), for a total of eight blood samples for each cow. On each blood sample total proteins and electrophoresis analysis were performed. On the data obtained, normally distributed (P<0·05, Kolmogorov-Smirnovs test), one-way Repeated Measure Analysis of Variance (ANOVA), was applied to evaluate the influence of different stages of gestation and lactation on the considered parameters. Results showed a significant effect on total proteins, α1-globulins, β-globulins, γ-globulins and albumin/globulin ratio. Most of the detected modifications were related to the transition from gestation to lactation, indicating that it is a period of great metabolic stress for cows. On the basis of the obtained results we can affirm that the pattern of serum protein fraction rn could give information about dehydration, plasma volume expansion and hepatic function occurring during the peripartum period in dairy cows.

Daily locomotor activity in five domestic animals

The aim of this study was to compare locomotor activity in five domestic animals. Five female Comisana sheep, five female Maltese goats, five female Bruna cows, and five female Thoroughbred horses were housed separately in a soundproof box equipped with an airflow system and five female Blue Vienna rabbits were individually housed in metallic cages. Animals were kept under a 12/12 Light/Dark cycle and indoor ambient temperature (16-20°C). To record activity, an Actiwatch-Mini ® was placed on each animal by means of a collar. Analysis by the cosinor procedure indicated a daily rhythm of locomotor activity in each of the five species studied. The main activity was observed during the photophase in sheep, goats, cows and horses, and during the scotophase in rabbits. The comparison among the species studied underlined a stable daily rhythm of locomotor activity in cows and horses, while sheep, goats and rabbits showed a weak daily rhythm of locomotor activity.

Blood gas analyses, ruminal and blood pH, urine and faecal pH in dairy cows during subacute ruminal acidosis

The aim of this study was to investigate the variations of some physiological parameters in dairy cows during subacute ruminal acidosis (SARA), a common important disorder of dairy cows that occurs in early lactation. pH changes in ruminal fluid collected by rumenocentesis were measured at ten farms stationed in different zones in the north of Italy. Additionally, the following parameters were measured: blood pH, faecal pH, urine pH, partial pressure of carbon dioxide oxygen, partial pressure of oxygen, bicarbonate level, base excess of extracellular fluid and oxygen content. Herds were divided into two groups according to their average ruminal pH: group A included farms with average ruminal pH > 5.8 (normal) and group B farms with average ruminal pH < 5.8 (acidosis). Unpaired Student’s t test was used to reveal statistical significances between the two groups. Ruminal pH changes due to pathogenesis can be diagnostic for SARA.