Influence of improved FcRn binding on the subcutaneous bioavailability of monoclonal antibodies in cynomolgus monkeys

Engineering monoclonal antibodies (mAbs) with improved binding to the neonatal Fc receptor (FcRn) is a strategy that can extend their in vivo half-life and slow their systemic clearance. Published reports have predominantly characterized the pharmacokinetics of mAbs after intravenous administration. Recently, studies in mice suggest FcRn may also play a role in affecting the subcutaneous bioavailability of mAbs. Herein, we examined whether five mAbs engineered with the T250Q/M428L Fc mutations that improved their FcRn interactions, and subsequently their in vivo pharmacokinetics after intravenous administration, had improved subcutaneous bioavailability compared with their wild-type counterparts in cynomolgus monkeys. Similar to the intravenous administration findings, the pharmacokinetic profiles of our variant mAbs after subcutaneous injection showed improved half-life or clearance. In contrast, a clear effect was not observed on the subcutaneous bioavailability. We expect that while FcRn may play a role in determining mAb subcutaneous bioavailability, multiple biopharmaceutical and physiological factors are likely to influence the success of engineering strategies aimed at targeting this pathway for improving bioavailability.     

Optimal conditions for blastogenic response of peripheral blood lymphocytes to T and B cell mitogens in cynomolgus monkeys

Optimal conditions for the mitogen-induced proliferation of T and B lymphocytes of cynomolgus monkeys were determined. The T cell mitogens concanavalin A and phytohemagglutinin, at concentrations of 1.25–10 μg/ml and 1.25–10 μg/ml, respectively, and the T and B cell mitogen pokeweed mitogen, at concentrations of 0.2–10 μg/ml, induced high lymphoproliferative responses, the average stimulation index (SI) being 34–93. Since suitable mitogens have not been reported for monkey B cells, we tested three types of lipopolysaccharide (LPS): two derived from Escherichia coli—one extracted with phenol and one extracted with trichloroacetic acid (TCA); and one derived from Salmonella typhimurium, extracted with phenol. All three LPS had a high mitogenic effect for monkey lymphocytes, with SI of 2.3–6.4. The highest response was observed for 25 μg/ml of Salmonella LPS, and the addition of trypsin to the culture augmented the proliferative response of low or non-responder lymphocytes. © 1994 Wiley-Liss, Inc.     

Effect of testosterone on epididymal proteins in castrated rhesus monkeys

The effect of testosterone on regulation of epididymal protein synthesis has been investigated in castrated rhesus monkeys (Macaca mulatta). The proteins in the treated monkeys were characterized using polyacrylamide gel electrophoresis (under nondenaturing and denaturing conditions) and electrofocusing. At least four distinct proteins have been shown to be synthesized by the monkey epididymis under testosterone influence. Two of these proteins were detected following two days of testosterone treatment while the other two proteins were detected after a six-day treatment period. None of these proteins was detectable in monkeys treated with estradiol for six days. Electrofocusing of epididymal cytosol proteins from untreated and testosterone-treated and castrated monkeys also confirmed the presence of four androgen-dependent proteins in this species. The isoelectric points of these proteins were shown to range between 5.8 and 6.4. The molecular weights of these proteins were found to vary between 47,500 and 66,000. The in vitro incorporation of ³H-labeled amino acids was markedly greater in the androgen-primed epididymis as compared with the control tissue.     

Two new species of night monkeys,genus Aotus (Cebidae,platyrrhini): A preliminary report on Aotus taxonomy

The nine allopatric species of Aotus recognized represent two natural groups distinguished by karyotype, color, and pelage patterns. Correlated with these group characters are reported differences in serum proteins and degrees of susceptibility or immunity to experimental infection with malarial parasites. The primitive gray-neck species group of Aotus contains A. brumbacki (new species), A. lemurinus (with subspecies lemurinus and griseimembra), A. trivirgatus, and A. vociferans. The derived red-neck group contains A. nancymai (new species), A. miconax. A. infulatus, and A. azarae (with subspecies azarae and boliviensis). Only the two new species are described but a key to the species and subspecies gives the diagnostic characters of each. The gray-neck group occurs almost entirely north of the Amazon, the red-neck group almost entirely south. The distributional exceptions are enclave populations resulting from river bend cutoffs. Formation of an enclave population of A. nancymai is discussed and available information on the biology of this species is reported.     

Influence of breeding site availability on the oviposition behaviour of Aedes aegypti

Despite the importance of the mosquito Aedes aegypti in thetransmission of arboviruses, such as yellow fever, Chikungunya fever and denguefever, some aspects of their behaviour remain unknown. In the present study, theoviposition behaviour of Ae. aegypti females that were exposed todifferent densities of breeding sites (2, 4, 8 and 16) was evaluated in laboratoryand semi-field conditions. The number of breeding sites that were used wasproportional to the number available, but tended towards stabilisation. Females usedfour-six breeding sites on average, with a maximum of 11. A high percentage of eggswas observed in the water, along with the presence of a breeding site termed“favourite”, which received at least 40% of the eggs. The results are discussed inecological, evolutionary and epidemiological approaches.     

Use of positive reinforcement training in the management of species for reproduction

Positive reinforcement training holds great potential for enhancing the management of species for reproduction. This paper reviews a wide range of animal training activities and resultant benefits which could have application to captive reproduction programs. Recognized benefits fall into several categories. Basic animal care can be improved through voluntary cooperation by the animals with veterinary procedures and routine husbandry activities. The quality and quantity of physiological data collected for research can be improved through voluntary cooperation by the animal subjects with sample collection. Positive social interaction and reproductive behavior can be increased and aggressive behavior reduced through employment of a specialized training regime called cooperative feeding. Good success has been reported with special training programs to facilitate introduction of new members into primate social groups, while mixed results have been obtained in efforts to use training to enhance maternal skills. Finally, a discussion of both direct and indirect enhancement of psychological well-being through the use of positive reinforcement training is presented. While positive reinforcement training techniques will not be useful in every situation, information developed to date strongly suggests that these techniques should be given serious consideration when developing comprehensive programs for the management of species for reproduction. © 1994 Wiley-Liss, Inc.     

Successful implementation of cooperative handling eliminates the need for restraint in a complex non-human primate disease model

Background Streptozotocin‐induced diabetic non‐human primates are used to study efficacy and safety of innovative immunosuppression after islet transplantation. We implemented a training program for medical management of a chronic disease state. Methods Cooperation with hand feeding and drinking, shifting, and limb presentation were trained utilizing predominately positive but also negative reinforcement in 52 animals compared with 28 macaques subjected to conventional physical and/or chemical restraint. The success and timing of behavior acquisition was evaluated in a representative subset of 14 animals. Results Over 90% of animals were successful in behavior acquisition. Programmatically this resulted in complete elimination of chair restraint and negligible requirement for sedation. About half of the trained animals had no‐to‐moderate thymic involution, indicative of a substantial reduction in stress. Conclusion Cooperative handling enhances animal well‐being. This contributes to validity of scientific results and eliminates model‐induced confounding that can obstruct interpretation of safety and efficacy data.     

Influence of training in the dark or light phase on physiologic and metabolic parameters of Wistar rats submitted to aerobic exercise

Light and dark phase training may influences rodents’ physiologic parameters because these animals have nocturnal habits. Thus, we verify the effects of the training in different photoperiods on metabolism and corporal composition of rats. Eighteen rats were divided into three groups – G1: non-trained; G2: trained in the light phase; G3: trained in the dark phase. Rats were allowed to swim for 60 min, five times per week during six weeks. Trained animals presented a smaller weight gain and fat percentage in carcass. Rats of G3 increased gastrocnemius relative weight. The adipocyte diameter of G3 rats was smaller than the other groups. The levels of the total cholesterol, low-density proteins, and triacylglycerols were decreased in animals of G2 while the glycemia was increased. Training in light phase provided more alterations in the blood biochemical profile while the training in the dark increased the gastrocnemius weight and decreased the diameter of the adipocyte.     

Effect of milking frequency and nutritional level on aspects of the health and welfare of dairy cows

The objective of this experiment was to investigate the effect of milking frequency and nutritional level on some aspects of animal health. Holstein-Friesian cows (n = 60) were assigned post calving to a factorial arrangement of treatments; twice a day (TAD) milking on a high or low nutritional level; once a day (OAD) milking on a high or low nutritional level. Milking characteristics were recorded daily. Blood samples to evaluate changes in the composition of the blood cells, milk leakage, udder tension and locomotory ability were measured on four occasions. Teat-ends were classified for hyperkeratosis (HK) monthly post partum. TAD had longer daily milking times (P < 0.001) compared with OAD cows. There was no effect of milking frequency or nutritional level on morning milking time, time to milk letdown or peak milk flow rate (P>0.05). High nutritional level cows had higher average flow-rates (P < 0.05) than low nutritional level cows. Neither milking frequency nor nutritional level affected HK (P>0.05). However, HK values were positively correlated with daily milking time for OAD cows for 6 months of lactation (P < 0.05). This correlation was significant (P < 0.01) for cows milked OAD on high nutrition during the peak lactation period. OAD cows had higher levels of milk leakage compared with TAD cows during the month of May (P < 0.01). Cows on high nutrition milked OAD showed higher udder firmness scores than cows milked once or twice daily at the low nutrition level in June and July (P < 0.05). OAD cows had higher locomotion scores compared with TAD cows (P < 0.001). Locomotion and udder firmness scores were significantly correlated for OAD in June (P < 0.05). OAD cows had lower blood lymphocyte counts, numerically higher counts of neutrophil and a higher monocyte count at peak lactation compared with TAD cows suggesting that OAD cows had altered immune responses. The increase in milk leakage, higher udder firmness and locomotion scores in conjunction with changes in blood cells, suggests that OAD milking may have caused some discomfort to the cows during peak lactation. A reduction in the concentrate input to OAD cows during this period could ameliorate this problem.     

Assessment of training effects on activity levels of alveolar macrophage in matured rats using chemiluminescent technique

Chemiluminescence responses have been used for the evaluation of phagocyte function. In this study, to evaluate effects of training started after maturation on pulmonary immunity, the activity levels of rat alveolar macrophages (AMs) were assessed as reactive oxygen species (ROS) generating capacity, measured by lucigenin‐ and luminol‐dependent chemiluminescence, using a parallel luminometer. One group of male Wistar rats started training at 11 weeks old and another group at 17 weeks old. The experimental period was 12 weeks, and about half of the rats were sacrificed after 6 weeks. The forced and voluntary exercises affect the mean levels of body weights and cell populations in the bronchoalveolar lavage fluid in younger animals; however, the voluntary exercise group in younger animals seemed to adapt after 12 weeks. By contrast, chemiluminescence responses in older rats observed after 6 weeks suggest that AMs are primed, and the maximum releasing activities of ROS are reduced. These changes in AM activity may be caused by the exercise and separation stresses and the rats may adapt to those stressors after 12 weeks. The chemiluminescent technique is thought to be useful to evaluate the changes of AM activity. Copyright © 2002 John Wiley & Sons, Ltd.     

Effects of oral functional training for nutritional improvement in Japanese older people requiring long-term care

Objectives: The aim of this study was to evaluate the effect of oral functional training and nutrient supplements to improve the nutrition of malnourished elderly people in a nursing home. Background: Malnutrition is a frequent problem in the elderly requiring long‐tem care; however, it is not clear whether oral functional training can be effective to improve nutrition. Subjects and methods: Fourteen subjects of 82 residents (mean age 85.7 ± 6.2 years) in a nursing home, who had a serum albumin level of ≤3.8 g/dl and understood the purpose of this study, were randomly divided into two groups of seven: one group served as the supplement group (mean age 87.0 ± 4.9 years) to which a high‐calorie and high‐protein diet was provided, and the other as the oral training plus supplement group (mean age 84.6 ± 10.1 years) to which oral functional training was given by a dental hygienist once a week as well as the above diet. Nutritional status was evaluated using serum biochemical values as indices at 4 months after the start of the intervention. Results: In the supplement group, serum albumin was 3.44 ± 0.36 g/dl at the start of the study (before intervention) and 3.24 ± 0.45 g/dl at 4 months after intervention. In the oral training plus supplement group, it was 3.56 ± 0.22 g/dl before intervention and significantly increased to 3.70 ± 0.33 g/dl after intervention (p < 0.05: Wilcoxon signed‐rank test). Conclusion: Nutritional supplements alone were not adequate, but with oral functional training to maintain and improve feeding function, nutritional improvement in the elderly could be observed.     

Variations in the timing of reinforcement as a training technique for foals (Equus caballus)

Abstract

Horses are used worldwide for a range of activities. Their usefulness and welfare in these pursuits are strongly influenced by their trainability, which may in turn be influenced by learning ability. Handling and riding horses can expose both handler and horse to a considerable risk of injury. This risk can be reduced by employing correct handling procedures that can facilitate learning in horses. As with all training, efficacy is influenced by consistency and timing. To determine the optimum timing of reinforcement, 16 unweaned naïve foals, that had previously undergone minimal human–animal interaction (i.e., not had a headcollar previously applied), of warmblood (WB; n = 6), thoroughbred (TB; n = 5) or warmblood x thoroughbred (WB x TB; n = 6) breeding were randomly assigned to three treatment groups for testing on ten training days at approximately 14-day intervals. Pressure applied to a headcollar via a lead rope was used as the stimulus for each foal to walk forward, and this was repeated until the foal had walked a distance of 8 m. The effects of three different latencies of negative reinforcement were evaluated by releasing the pressure either immediately: as the first foreleg step commenced (Treatment 1); when the second step of the forelegs was completed (Treatment 2) or when the fourth step of the forelegs was completed (Treatment 3). Each foal's rate of learning was measured by the proportion of correct responses relative to the total number of responses performed. Behavioral responses exhibited (rears, strikes, head shakes, falls, sideways movement and hops) and the steps taken over the distance were also recorded.

Initially, the foals undergoing Treatment 1 appeared to learn more quickly than those foals undergoing Treatments 2 and 3, suggesting that Treatment 1 was associated with the greatest compliance and the quickest learning. However, the foals undergoing Treatment 3 ultimately achieved significantly (p < 0.001) more correct responses, suggesting that the longer delay of reinforcement (i.e., the longer duration of aversive stimulus) may enhance learning via the negative reinforcement inherent in lead training in foals. While some conflict behaviors were shown in all treatment groups, most were exhibited on training day 2. This was reflected in the analysis of composite behaviors performed, with training days 1 and 2 being different (p < 0.001) from training day 3, and training days 1–3 being different (p < 0.001) from training days 4–10. These changes indicate that learning occurred in all treatment groups.

The foals used in this study were sired by five different stallions. While the foals sired by stallion 2 (WB) performed significantly (p < 0.001) more correct responses, those foals sired by WB stallions were significantly (p < 0.001) less likely to perform correct responses when compared with those foals of TB or WB x TB breeding. Colts were significantly (p < 0.001) more likely to perform correct responses than were fillies.     

Effect of Breed,Cage Type,and Reproductive Phase on Fecal Corticosterone Levels in Doe Rabbits

Fecal corticosterone concentration (FCC) is increasingly being used as a noninvasive indicator of stress in assessment of nonhuman animal welfare. The aim of this study was to evaluate effects of breed, cage type, reproductive phase, and their interactions on FCC levels in doe rabbits. A total of 252 doe rabbits were randomly assigned to 2 groups. Does were individually housed in either standard dimension cages (SC) or in cages with a volume more than double that of the SC. Bigger cages (BC) were equipped with a plastic foot mat. Breed, cage type, and reproductive phase significantly affected FCC. New Zealand hybrids showed higher FCCs (p < .001) when compared witho the autochthonous breed (27.77 ± 0.47 vs. 24.61 ± 0.36 pg g^?1, respectively). Increased cage size coupled with a plastic foot mat resulted in a significant decrease in doe FCC. The highest FCCs were detected at partum (BC: 30.40 ± 0.81 pg g^?1; SC: 33.36 ± 0.86 pg g^?1; p ≤ .05), followed by postweaning (BC: 25.09 ± 0.95 pg g^?1; SC: 27.63 ± 0.95 pg g^?1; p ≤ .05). These results support the hypothesis that measurement of FCC provides a useful indicator of chronic stress in doe rabbits.     

Effect of exercise training on total daily physical activity in elderly humans

This study examined the effect of 12 weeks of exercise training on daily physical activity in elderly humans. Training consisted of a weekly group session and an individual session with cardio- and weight-stack machines. A group of 15 subjects served as the exercise group [EXER mean age 59 (SD 4) years], and 7 subjects as the controls [CONT mean age 57 (SD 3) years]. Physical activity and physical fitness were measured before the start of training (T), at week 6 and week 12 (T0, T6, T12 respectively) in EXER, and at T0 and T12 in CONT. Physical activity over 14 days was measured using a tri-axial accelerometer and physical fitness was measured during an incremental exercise test. At T12, mean maximal power output had significantly increased in EXER compared to CONT 8 (SD 12) vs -5 (SD 9) W; P < 0.02] and mean submaximal heart rate (at 100 W) had reduced [-10 (SD 7) vs -2 (SD 6) beats x min(-1); P < 0.05]. No differences or changes in physical activity were observed between EXER and CONT. At T6, physical activity on training days was significantly higher than on non-training days (P < 0.001). When the accelerometer output of the training session was subtracted from the accelerometer output on training days, at T12 non-training physical activity was significantly lower than on non-training days (P < 0.004). Accelerometer output of the individual training session at T12 had significantly increased compared to T6 (P < 0.05), whereas, accelerometer output of the group training session had remained unchanged. In conclusion, in elderly subjects an exercise training programme of moderate intensity resulted in an improved physical fitness but had no effect on total daily physical activity. Training activity was compensated for by a decrease in non-training physical activity.     

Effect of increasing trypsin concentrations on seminal coagulum dissolution and sperm parameters in spider monkeys (Ateles geoffroyi)

Seminal coagulum formation in spider monkeys (Ateles geoffroyi) interferes with the efficient recovery and evaluation of spermatozoa. The main objective was to assess the effect of increasing concentrations of trypsin on dissolution of seminal coagulum and spermatic parameters. Seminal coagulum was incubated at 37 °C without trypsin or in the presence of increasing trypsin concentrations (0.1%, 1.0%, and 5.0%). For each sample, coagulum dissolution time was measured, and sperm concentration, viability, motility, and morphology were evaluated using light microscopy and/or transmission electronic microscopy (TEM). Trypsin concentrations of 1.0% and 5.0% more rapidly liquefied seminal coagulum, averaging 32 and 21 min, respectively, compared with nontrypsinized controls, with maintenance of greater sperm viability (70.8% and 72.5%, respectively). Coagulum treated with 1.0% trypsin and the liquid ejaculate fraction averaged higher sperm motility (40.1% and 55.6%, respectively) than control samples, and both 1.0% and 5.0% trypsin treatment allowed recovery of increased numbers of motile spermatozoa. There was greater sperm fragmentation at the head and midpiece level after treatment with 1.0% and 5.0% trypsin (55.8% and 55.9%); however, the percentage of normal morphology in structurally intact spermatozoa did not differ relative to controls. With transmission electronic microscopy imaging, there were similar percentages of spermatozoa with plasma membrane swelling in the midpiece and acrosomal regions in trypsin-treated samples and controls. In conclusion, trypsin treatment of spider monkey seminal coagulum exerted a concentration-dependent effect on dissolution time and various spermatic parameters. Higher trypsin concentrations caused more rapid liquefaction of coagulum and recovery of greater numbers of motile spermatozoa, but may adversely affect fragmentation of spermatozoa and could compromise sperm function and cryopreservation potential.     

Capillary affinity gel electrophoresis: new technique for specific recognition of DNA sequence and the mutation detection on DNA

The present state of studies on capillary affinity gel electrophoresis, which is a new technique for the specific recognition of a target DNA sequence, is reviewed. This article includes the principle, theory, methods, and applications of this technology. The great potential of capillary affinity gel electrophoresis for the sequence-specific recognition of DNA and the detection of mutations in specific genes is illustrated.     

Development of an Optimal Diaphragmatic Hernia Rabbit Model for Pediatric Thoracoscopic Training

Our objectives were to standarize the procedure needed to reproduce a similar surgicalscene which a pediatric surgeon would face on repairing a Bochdalek hernia in newborns andto define the optimal time period for hernia development that achieve a realistic surgicalscenario with minimimal animal suffering. Twenty New Zealand white rabbits weighing 3–3.5kg were divided into four groups depending on the time frame since hernia creation tothoracoscopic repair: 48 h, 72 h, 96 h and 30 days. Bochdalek trigono was identified andprocedures for hernia creation and thoracoscopic repair were standarized. Blood wascollected for hematology (red blood cells, white blood cells, platelets, hemoglobin andhematocrit), biochemistry (blood urea nitrogen, creatinine, alanine aminotransferase,aspartate aminotransferase, lactate dehydrogenase and creatine kinase) and gas analysis(arterial blood pH, partial pressure of oxygen, partial pressure of carbón dioxide, oxygensaturation and bicarbonate) at baseline and before the surgial repairment. Glucocorticoidmetabolites concentration in faeces was measured. Thoracoscopy video recordings wereevaluated by six pediatric surgeons and rated from 0 to 10 according to similarities withcongenital diaphragmatic hernia in newborn and with its thoracoscopic approach.Statistical methods included the analysis of variance, and comparisons between groups werefollowed by a post-hoc Tukey’s test. Fourty -eight h showed to be the optimal time frameto obtain a diaphragmatic hernia similar to newborn scenario from a surgical point of viewwith minimal stress for the animals.     

Effect of running training on uncoupling protein mRNA expression in rat brown adipose tissue

The effect was investigated of endurance training on the expression of uncoupling protein (UCP) mRNA in brown adipose tissue (BAT) of rats. The exercised rats were trained on a rodent treadmill for 5 days per week and a total of 9 weeks. After the training programme, a marked decrease in BAT mass was found in terms of weight or weight per unit body weight; there was a corresponding decrease in DNA content and a downward trend in RNA and glycogen levels. The UCP mRNA was present at a markedly decreased level in BAT of trained animals. In consideration of the reduced levels of mRNAs for hormone-sensitive lipase and acylCoA synthetase, the brown adipose tissue investigated appeared to be in a relatively atrophied and thermogenically quiescent state.