Evaluation of Recombinant Canine Granulocyte Colony-Stimulating Factor as an Inducer of Granulopoiesis: A Pilot Study

Five healthy young adult dogs were given recombinant canine granulocyte colony-stimulating factor (rcG-CSF) at a dosage of 5 micrograms/kg/day subcutaneously for 4 weeks to evaluate the effect on complete blood cell counts. The mean neutrophil counts +/- standard deviation (SD) increased significantly (P less than 0.01) from 6,537/microliters +/- 1,726 (range, 4,950-9,512/microliters) to 26,330/microliters +/- 7,066 (range, 15,368-35,785/microliters) within 24 hours after the first injection of rcG-CSF. Mean monocyte counts +/- SD were significantly increased (P less than 0.05) from baseline values of 751/microliters +/- 168 (range, 444-891/microliters) to 2,514/microliters +/- 878 (range, 1,740-3,752/microliters) on day 5 of rcG-CSF administration. Mean neutrophil and monocyte counts (+/- SD) continued to increase reaching a maximum of 72,125/microliters +/- 15,073 (range, 50,915-96,278/microliters) and 3,972/microliters +/- 2,621 (range, 685-8,030/microliters), respectively by day 19. These increased neutrophil and monocyte counts were maintained until the administration of rcG-CSF was stopped. Blood counts returned to normal within 5 days after discontinuing the rcG-CSF. One week after discontinuing treatment, rcG-CSF was started again at 5 micrograms/kg/day subcutaneously. Within 48 hours following administration of rcG-CSF, mean neutrophil counts +/- SD increased from 5,860/microliters +/- 1,819 (range, 3,720-8,650/microliters) to 57,444/microliters +/- 8,173 (range, 43,983-68,278/microliters). Myeloid:erythroid ratios increased from a mean of 1.63:1 on day 1 prior to administration of rcG-CSF to 3.3:1 on day 10 in three dogs for which bone marrow samples were evaluated. Recombinant canine G-CSF did not cause clinically significant toxicosis in any of the dogs.     

Use of Recombinant Canine Granulocyte Colony-Stimulating Factor to Decrease Myelosuppression Associated with the Administration of Mitoxantrone in the Dog

Ten dogs were given mitoxantrone at a dose of 5 mg/m2 body surface area intravenously. Recombinant canine granulocyte colony-stimulating factor (rcG-CSF) was administered subcutaneously daily for 20 days after an infusion of mitoxantrone in five of these dogs to determine the effect of the hematopoietic growth factor on the duration and severity of myelosuppression. The median neutrophil counts dropped below normal (less than 3,000/uL) for 2 days in the dogs that received rcG-CSF, and for 5 days in the dogs that received only mitoxantrone. Four of five dogs not treated with rcG-CSF and none of those receiving rcG-CSF developed serious neutropenia (less than 1,500/uL). The neutrophil counts were significantly (P less than 0.05) higher in the rcG-CSF treated dogs at all time points except before the administration of the colony-stimulating factor, and the sixth day after the mitoxantrone was administered. These findings demonstrate that rcG-CSF is capable of reducing the duration and severity of mitoxantrone-induced myelosuppression.     

犬转移因子对犬淋巴细胞体外增殖的影响

无菌采取犬抗凝血,分离并制备不同浓度外周血单个核细胞(PBMC)悬液。应用正交试验,测定植物血凝素,犊牛血清对不同浓度PBMC增殖的影响。用 MTT法测定不同浓度犬转移因子(TF)对 PBMC 增殖的影响。结果表明,当 PBMC 为 5×109 个/L,植物血凝素为 12. 5 mg/L, 犊牛血清为100 mL/L,PBMC 增殖效果最佳。TF 浓度在0.39 g/L~100 g/L时,对 PBMC转化增殖均有促进作用,在浓度为 1.56 g/L时促淋转效果最佳。且TF单独作用于 PBMC的效应明显优于PHA P的协同作用。TF对 PB MC有良好的刺激增殖效果。     

Thrombokinetic Studies in Normal and Factor VIII-deficient Canine Plasmas

Thrombokinetograms are graphic depictions of the optical changes occurring in plasma during the clotting process and provide information, not only on the time required for clotting to begin, but also on the way in which the clot forms. We studied thrombokinetic profiles in plasmas from normal dogs, and dogs with varying degrees of factor VIII deficiency. Clotting was induced through intrinsic, extrinsic and common coagulation pathways [activated partial thromboplastin time, prothrombin time and thrombin time, respectively].

The thrombokinetograms for the various clotting tests were qualitatively similar in normal canine plasmas. After activation of the clotting system there was a period in which no change in optical density occurred. This period was represented by the left base line and corresponded to the duration of the clotting time. When fibrin production commenced there was a rapid increase in the rate of optical density change (ΔOD) to a maximum (V_maxΔOD) in time t₁. This was followed by a more gradual reduction in ΔOD in time t₂.

The activated partial thromboplastin time thrombokinetograms for von Willebrand's disease plasmas were characterized by a reduced V_maxΔOD and prolonged t₁. In severe hemophilic plasma [factor VIII coagulant (F VIII:C)<1% of normal] there was a very slow increase in ΔOD following a prolonged left baseline. The V_maxΔOD, t₁ and t₂ could not be determined since a peak was not attained in one minute. The prothrombin and thrombin time thrombokinetograms for von Willebrand's disease plasmas were normal. The prothrombin time thrombokinetogram for hemophilic plasma had a 2X normal V_maxΔOD possibly related to the relatively high fibrinogen concentration of this plasma compared to the normal.

Changes in thrombokinetogram profiles may be of value in studying mild to moderate clotting factor deficiencies particularly where the clotting times are not markedly prolonged.

     

Peripheral Blood Neutrophil Function and Lymphocyte Subpopulations in Cycling Mares

The purpose of this study was to evaluate different parameters of the immune status in the mare, during the follicular and the luteal phases of the oestrous cycle, in two consecutive years. Functional competence of peripheral blood neutrophils, such as chemotaxis, phagocytosis and oxidative burst was assessed under physiological cyclic conditions (Exp. I). In the second year of this study (Exp. II), besides peripheral blood neutrophil phagocytosis and oxidative burst analysis, circulating lymphocyte subsets were also characterized. The reproductive status in a total of 17 adult cycling mares was evaluated by ultrasonography and further confirmed by plasma progesterone levels. Chemotaxis tests were performed using porous membranes inserted in transwell chambers. Lipopolysaccharide (LPS) from Escherichia coli and N‐formyl‐Met‐Leu‐Phe (fMLP) were used as chemoattractants. Measurement of phagocytosis and oxidative burst in blood neutrophils were assessed by flow cytometry using commercially available kits. Quantification of T‐lymphocyte subsets was assessed by indirect immunofluorescence staining after incubation with monoclonal antibodies specific for CD2, CD3, CD4 and CD8 cell markers by flow cytometry. Natural killer cells and B cells were estimated mathematically. No significant difference was found in migration, phagocytosis and oxidative burst at either phase of the oestrous cycle. Statistical analysis of total white blood cell counts also showed no significant difference between either phase of the oestrous cycle, although there was a tendency for blood neutrophils to increase in number under the progesterone influence (p = 0.09). Lymphocytic subpopulations did not differ throughout the oestrous cycle. Overall, our results suggest that luteal and follicular phases in cycling mares may not influence the immune status of the mare.     

两种转移因子对犬外周血T细胞亚群的影响

按标准方法分别提取制备了猪和犬的转移因子，选择杂种牧羊犬为试验犬，用流式细胞术检测了试验犬注射猪、犬转移因子前和注射后不同时间T细胞亚群的变化情况。结果表明，同种、异种转移因子均可提高试验犬外周血淋巴细胞数量；CD4/CD8比值变化分析结果证明，外周血增加的淋巴细胞主要为CD4+ T细胞；试验犬注射同种、异种转移因子后20 d内，CD4+ T细胞持续增加；同种转移因子促进试验犬外周血CD4+ T细胞上升的作用强于异种转移因子。     

苷肽注射液对犬外周血CD 4、CD 8比值的影响

以20只健康犬为研究对象,随机分为四组,每组5只,分别注射苷肽注射液、苷肽注射液和犬五联疫苗、犬五联疫苗、生理盐水,各组犬于注射的第0、2、7、15天采血,检测不同组之间的CD 4＋和CD 8＋比值变化规律。结果表明：苷肽注射液单独应用和与疫苗同时应用,在注射的第2天就可以显著提高CD4＋、CD8＋比值,到第7天时达到最高值。并且发现,注射苷肽注射液同时注射犬五联疫苗组和单独注射苷肽注射液组CD4＋、CD8＋比值显著高于单独注射犬五联疫苗组（P〈0.05）,这说明苷肽注射液具有增强免疫的作用。     

The Effect of Intercondylar Notchplasty on the Normal Canine Stifle

This study evaluated the effect of notchplasty (enlargement of the intercondylar fossa) in stable and unstable canine stifles. Bilateral notchplasty and unilateral cranial cruciate ligament (CrCL) transection were performed in 6 dogs. Exercise, consisting of walking 1.5 miles three times a week, began 1 month after surgery and continued until euthanasia 6 months after surgery. Evaluation methods included orthopedic examination, serial radiographs, thin section radiography, histopathology, and gross pathology. Notchplasty in the stable stifle did not cause lameness beyond 3 weeks, joint instability, or degenerative joint disease. In the stable stifle, smooth resurfacing of the notchplasty site with fibrous and osseous tissue occurred. Stifles with notchplasty and CrCL transection exhibited persistent lameness, instability, and degenerative joint disease. In CrCL deficient stifles osteophytes formed within the notchplasty site, resulting in a rough surface. Our observations indicated significant refilling in notchplasties of both stable and unstable stifles ( P <.05). However, the intercondylar fossa (ICF) width 6 months after notchplasty was significantly smaller in unstable stifles compared with stable stifles ( P <.05) indicating that greater refilling of the notchplasties occurred in the unstable stifles. In clinical cases, notchplasty should be larger than the desired final result to accomodate the partial refilling that occurs even in stable stifles.     

Oral Mucosa Bleeding Times of Normal Cats and Cats with Chediak-Higashi Syndrome or Hageman Trait (Factor XII Deficiency)

A commercially available, disposable blade in a spring-loaded cassette was used to measure oral mucosa bleeding times (OMBT) of ketamine/acepromazine-anesthetized cats. The OMBT were determined in cats homozygous for Chediak-Higashi syndrome (CHS, n = 7), cats heterozygous for CHS (n = 6), and cats homozygous for Hageman factor (factor XII) deficiency (n = 5). In addition, OMBT were determined in three groups of normal cats: random-source cats (n = 14), inbred normal relatives of the cats with CHS (n = 7), and inbred normal relatives of Hageman factor deficient cats (n = 9). No significant differences were found in the OMBT of the three groups of normal cats. The mean OMBT for all 30 normal cats was 1.9 minutes +/- 0.5 minutes s.d. Compared to the normal cats, those homozygous for CHS had significantly prolonged OMBT (14.1 +/- 3.3 minutes; p < 0.05). The mean OMBT of cats heterozygous for CHS (2.6 +/- 0.8 minutes) was also significantly longer than the OMBT of the combined normal group. The mean OMBT of the CHS heterozygotes, however, was not significantly longer than that of their normal relatives (OMBT = 1.8 +/- 0.5 minutes), probably because of the low number of cats in this subgroup of normals. As expected, the OMBT of cats homozygous for Hageman factor deficiency (2.3 +/- 0.3 minutes) were not significantly prolonged.     

Peripheral and Central Venous Blood Glucose Concentrations in Dogs and Cats with Acute Arterial Thromboembolism

Background

Acute limb paralysis because of arterial thromboembolism (ATE) occurs in cats and less commonly in dogs. ATE is diagnosed based on physical examination findings and, occasionally, advanced imaging.

Hypothesis/Objectives

Peripheral, affected limb venous glucose concentration is decreased in ATE, whereas its systemic concentration is within or above reference interval.

Animals

Client‐owned cats and dogs were divided into 3 respective groups: acute limb paralysis because of ATE (22 cats and 9 dogs); acute limb paralysis secondary to orthopedic or neurologic conditions (nonambulatory controls; 10 cats and 11 dogs); ambulatory animals presented because of various diseases (ambulatory controls; 10 cats and 9 dogs).

Methods

Prospective observational, clinical study. Systemic and local (affected limb) blood glucose concentrations were measured. Their absolute and relative differences (ΔGlu and %ΔGlu, respectively) were compared among groups.

Results

ΔGlu and %ΔGlu were significantly higher in the ATE cats and dogs groups, compared to both of their respective controls (P < .0001 and P < .001, respectively). No significant differences were observed between the control groups. Receiver operator characteristics analysis of ΔGlu and %ΔGlu as predictors of ATE had area under the curve of 0.96 and 0.99 in cats, respectively, and 1.00 and 1.00, in dogs, respectively. ΔGlu cutoffs of 30 mg/dL and 16 mg/dL, in cats and dogs, respectively, corresponded to sensitivity and specificity of 100% and 90% in cats, respectively, and 100% in dogs.

Conclusions and Clinical Importance

ΔGlu and %ΔGlu are accurate, readily available, diagnostic markers of acute ATE in paralyzed cats and dogs.     

锌对雏鸭外周血T-淋巴细胞的影响

15 0只 1日龄天府肉鸭健雏随机分为 3组 ,分别喂给缺锌 (每千克日粮含 Zn 2 2 .9mg)、对照 (每千克日粮含 Zn10 0 mg)和锌中毒 (每 kg日粮含 Zn130 0 mg)日粮 7周或 4周 ,以酸性 - α-醋酸萘酯酶 (ANAE)染色法观测外周血 T-淋巴细胞的动态变化。结果锌缺乏组和锌中毒组雏鸭外周血 T-淋巴细胞的 ANAE阳性率显著低于对照组 (P<0 .0 1) ,表明锌缺乏或锌中毒可抑制 T-淋巴细胞的生成 ,降低其在外周血中的数量。本试验还对锌缺乏和锌中毒引起的外周血 T-淋巴细胞减少的机理进行了探讨     

女贞枸杞散预防断奶仔猪腹泻效果和对血液白细胞数的影响

为探讨以女贞子、枸杞子、穿心莲、大青叶等多味中药组成的女贞枸杞散对断奶仔猪腹泻的预防效果和对外周血各类白细胞数量的影响。根据体重相近、公母各半的原则选用80头体重为10kg左右的杜长大三元杂交断奶仔猪,随机分4组,分别为试验Ⅰ组(日粮中加0.4%女贞枸杞散),Ⅱ组(添加0.6%),Ⅲ组(添加0.8%)和对照组(普通日粮)。详细记录仔猪腹泻情况,计算腹泻率、腹泻频率、腹泻指数等。试验结束后采血,测定各类白细胞数量,结果表明,女贞枸杞散能有效地降低断奶仔猪的腹泻率,改善仔猪的生长性能,促进血液中的白细胞尤其是淋巴细胞的的生成,提高机体免疫力。     

免疫增强剂TMFN对犬外周血CD4/CD8比值的影响

采用检测犬外周血 CD4和 CD8比值变化的方法 ,观察犬注射免疫增强剂 TMFN和犬疫苗后 ,不同组之间的 CD4和 CD8比值 ,以证明该免疫增强剂对犬免疫系统的影响。结果表明 ,该免疫增强剂能够提高正常犬的细胞免疫水平 ,在犬注射疫苗的同时应用该免疫增强剂 ,可显著提高疫苗的免疫效力     

锌中毒对雏鸡外周血T-淋巴细胞的影响

200只1日龄艾维菌肉鸡随机分为4组，即对照组（每千克日粮含锌100mg）、锌中毒Ⅰ组（每千克日粮含锌1500mg）、锌中毒Ⅱ组（每千克日粮含锌2000mg）、锌中毒Ⅲ组（每千克日粮含锌2500mg），每组均饲喂日粮7周。以流式细胞术和酸性α-醋酸萘酯醇（ANAE）染色法观测外周血T-淋巴细胞的动态变化。结果，3个锌中毒组7周龄时，外周血T-淋巴细胞ANAE阳性率显著低于对照组（P〈0．05或P〈0．01），同时3个锌中毒组间比较也呈显著差异（P〈0．05或P〈0．01）。与对照组比较，锌中毒Ⅲ组2至7周龄和锌中毒Ⅱ组6、7周龄CD4^＋T淋巴细胞数量减少，锌中毒Ⅱ组、Ⅲ组2至4周龄和锌中毒Ⅰ组、Ⅲ组7周龄CD8^＋T淋巴细胞数量降低，CD4^＋／CD8^＋比值3个锌中毒组2、4周龄升高，6、7周龄降低。由此表明，锌中毒可抑制T-淋巴细胞的生成，降低其在外周血中的数量。     

铜中毒对雏鸡外周血T-淋巴细胞的影响

180只1日龄艾维茵肉鸡健雏随机分为3组，分别饲喂对照（每千克日粮含铜11．97mg）和铜中毒（每千克日粮舍铜650mg，铜中毒Ⅰ组；每千克日粮含铜850mg，铜中毒Ⅱ组）日粮6周，以流式细胞术和酸性-α-醋酸萘酯酶（ANAE）染色的方法观测外周血T-淋巴细胞的动态变化。2个铜中毒组1～6周龄外周血T-淋巴细胞的ANAE阳性率均显著低于对照组（P〈0．05或P〈0．01），同时2个铜中毒组间比较也呈显著差异（P〈0．05或P〈0．01）。与对照组比较，2个铜中毒组2～6周龄CD4^＋T淋巴细胞数量均显著降低（P〈0．05或P〈0．01），CD8^＋T淋巴细胞数量变化不明显（P〉0．05），CD4^＋／CD8^＋比值降低（P〈0．05）。结果表明，铜中毒可抑制T-淋巴细胞的生成，降低其在外周血中的数量。     

Serum Progesterone Concentrations and Blood Neutrophil Alkaline Phosphatase Activity in Cows Following Normal and Abnormal Parturition

Serum progesterone concentrations and alkaline phosphatase (ALP) activity of blood neutrophils were determined in 3 groups of cows (n = 5 each) on days 1 and 2 and then at 3-day intervals up to 32 days post-partum. Group I cows had a normal delivery, Group II cows had dexamethasone-induced parturition and Group III cows were subjected to a caesarian section. All cows in Group III and 2 cows in Group II retained their fetal membranes. Mean serum progesterone concentrations declined the second day after calving (to < 0.67 ng/ml) and remained at low levels (< 0.54 ng/ml) throughout the observation period, except for the values in Group III, which were elevated on day 16 (0.94 ng/ml), declined again on day 26 (0.46 ng/ml) and peaked (1.05 ng/ml) on day 32 portpartum. Significant (P < 0.01) differences were found between serum progesterone concentrations on day 1 and on each of the other sampling days in Groups I and III. Day X parturition group interaction was significant (P < 0.05) for the progesterone concentrations. No significant differences were found between the overall means of ALP activity of blood neutrophils in the 3 parturition groups nor between days of the experiment. No significant correlation was found between serum progesterone concentrations and ALP activity values of blood PMN during the first 32 days post-partum. Inhalt: Serum Progesteron Konzentrat und Aktivität der alkalischen Phosphatase in neutrophilen Blutzellen bei Küken mit normalem und abnormalen Geburtsverlauf Serum-Progesteronkonzentrationen und alkalische Phosphatase- (ALP) Aktivität von neutrophilen Granulozysten, nach der Abkalbung am Tag 1 und 2 und dann im Abstand von 3 Tagen bis zum 32. Tag p.p. bei Kühen untersucht, die in drei Untersuchungsgruppen (n = 5) eingeteilt waren. Gruppe I kalbte normal ab, in Gruppe II wurde die Geburt durch Dexamethason eingeleitet, und Kälber der Gruppe III wurden via Kaiserschnitt gewonnen. Alle Tiere der Gruppe III und 2 Kühe der Gruppe II wiesen Nachgeburtsverhaltungen auf Der mittlere Serum-Progesterongehalt sank am 2. Tag p.p. auf < 0,67 ng/ml ab und blieb auf niederigem Niveau (< 0,54 ng/ml) während der gesamten Untersuchungs-periode. Lediglich in Gruppe III, mit erhöhten Werten (0,94 ng/ml) an Tag 16 p.p., sanken die Werte wieder an Tag 26 p.p. (0,46 ng/ml) und erreichten einen Maximalwert an Tag 32 p.p. mit 1,05 ng/ml. Signifikante (P 0,01) Unterschiede in der Serum-Progesteronkonrentration wurden zwischen Tag 1 p.p. und allen anderen Untersuchungstagen in den Gruppen I und III gefunden. Interaktionen zwischen den Gruppen für Tag X waren signifikant (P 0,05). Keine signifikanten Unterschiede wurden für den Gesamtmittelwert der ALP-Aktivität in den neutrophilen Granulorysten zwischen den Gruppen und im Vergleich der Untersuchungstage gefunden. Es wurden auch keine signifikanten Korrelationen zwischen den Serum-Progesteronkonzentrationen und der ALP-Aktivität der PMN während der ersten 32 Tage p.p. nachgewiesen.     

The Effect of Storage on Ammonia Concentration in Canine Packed Red Blood Cells.

Objective: To determine the effect of storage on ammonia concentration in canine packed red blood cell (pRBC) units.
Design: In vitro and in vivo study.
Setting: University Veterinary Teaching Hospital.
Interventions: Ammonia concentration was measured in 7 units of canine pRBC prepared in citrate-phosphate-dextrose (CPD) and Adsol^a on Days 1 and 35 of storage. Ammonia was measured in 4 additional units of canine pRBC on Days 0, 7, 14, 21, 28, and 35. Plasma ammonia was also determined in 5 anemic dogs receiving pRBC.
Measurements and Main Results: Ammonia concentration increased from 73 ± 15 mmol/L (mean ± SD) on Day 1 to 800 ± 275 mmpl/L on Day (p<0.001). When measured every 7 days in 4 units of canine pRBC, ammonia concentration increased from 23 ± 8 mmol/L on Day 0 to 179 ± 13 mmol/L (Day 7), 276 ± 56 mmol/L (Day 14). 383 ± 47 mmol/L (Day21), 466 ± 30 mmol/L (Day 28), and 562 ± 27 mmol/L (Day 35) (p<0.05 for all comparisons). In a preliminary study, plasma ammonia concentration measured in blood samples from 5 anemic dogs without primary liver disease immediately before and after transfusion with 5–10 ml/kg of stored pRBC remained in the normal reference range.
Conclusions: The ammonia concentration in stored canine pRBC increased markedly with time. In this preliminary study, ammonia concentrations in dogs without primary liver disease did not increase above the reference range after transfusion with pRBC.     

Effect of Anticoagulants and Sampling Time on Results of Progesterone Determination in Canine Blood Samples

The concentration of plasma progesterone was measured by ELISA, in serum and samples prepared with three different anticoagulant agents – namely ethylenediaminetetraacetic acid (EDTA), heparine and sodium fluoride oxalate potassium(NaFK). Forty clinically healthy bitches were selected based on the signs of pro‐oestrus or oestrus. Values of progesterone concentration were significantly higher in serum than in EDTA‐plasma (p < 0.0005); heparin‐plasma (p < 0.05) and NaFK‐plasma (p < 0.005). During pro‐oestrus and oestrus until the time of ovulation, progesterone exhibited a conspicuous and statistically verified diurnal pattern (p < 0.05), its serum concentration being higher during 6.00–7.00 p.m. than 8.00–9.00 a.m. By the time of ovulation tendency of higher p.m. progesterone level reverses and from this point on the a.m. progesterone concentration is higher. The results of these experiments indicate that the concentration of canine progesterone assayed with ELISA may be affected by the time of collection and the method of preservation used.