The effect of inhibition of heat-shock proteins on thiram-induced tibial dyschondroplasia.

Tibial dyschondroplasia (TD) is a skeletal abnormality that can cause economic losses and animal welfare concerns. Thiram-induced TD is characterized by enlarged, unvascularized growth plates, low levels of the vascular endothelial growth factor receptor Flk-1, abnormal chondrocyte differentiation, and lameness. Recently we reported the involvement of heat-shock protein 90 (Hsp90) in chondrocyte differentiation and growth-plate vascularization. Inhibition of Hsp90 activity in thiram-induced TD resulted in increased Flk-1 levels, re-instated normal growth-plate angiogenesis and morphology, and abrogated lameness. In the present study, we evaluated the efficacy of various concentrations of 17-(dimethylaminoethylamino)-17-demethoxygeldanamycin (17-DMAG), an inhibitor of Hsp90 activity, in preventing growth-plate histopathology and lameness in TD-affected chicks. Low doses of 17-DMAG (2 injections, each of 100 or 300 µg) did not prevent TD development even though Flk-1 levels were restored, which suggests that Flk-1 is not the only rate-limiting factor in growth-plate angiogenesis. High doses of 17-DMAG (2 injections, each of 600 or 900 µg) prevented BW loss, decreased the TD score, reduced lesion width, restored proper chondrocyte differentiation, increased blood vessel invasion, and eliminated lameness. To assess the specificity of Hsp90, we evaluated the efficacy of the flavonoid quercetin, an inhibitor of Hsp70 synthesis, in preventing TD development; it decreased Hsp70 levels but not those of Hsp90 in the control growth plates and prevented upregulation of Hsp70 in the TD-affected growth plates. Dietary quercetin (at 100 or 500 ppm) did not prevent the hypoxia that is characteristic of the TD-affected growth plate or development of thiram-induced TD and lameness. The present results demonstrate the specificity and the major role of Hsp90 in chondrocyte differentiation and growth-plate vascularization. In contrast to the anti-angiogenic effect of 17-DMAG observed in mammals, inhibition of Hsp90 activity in the unvascularized TD-affected growth plates resulted in activation of the angiogenic switch and restored normal growth-plate morphology.

Hypoxia, hypoxia-inducible factor-1alpha (HIF-1alpha), and heat-shock proteins in tibial dyschondroplasia.

Tibial dyschondroplasia (TD) is one of the most prevalent skeletal abnormalities in avian species; it causes economic losses and is an animal welfare problem. It has been hypothesized that the absence of vasculature in the lesion of the TD growth plates at the ends of the long bones is involved in the etiology of the disease. We evaluated the hypoxia status of normal and thiram-induced TD growth plates by immunostaining the protein adducts after pimonidazole hydrochloride administration. In addition, we evaluated the expression of hypoxia-inducible factor-1alpha (HIF-1alpha), the major regulator of the hypoxic response that is essential for chondrogenesis, and that of heat-shock proteins (Hsp) downstream from HIF-1alpha. We demonstrated that, in contrast to the normal growth plates, those afflicted by TD were hypoxic. A major increase in hypoxia was observed in the proliferative, hypertrophic, and calcified zones. In the normal growth plate, HIF-1alpha was expressed in chondrocytes of the articular cartilage and of the maturation zone, whereas in cases of TD, HIF-1alpha was also expressed in chondrocytes below the lesion. The expression level of HIF-1alpha was related to the severity of the disease, but was independent of its cause; the same pattern of expression was observed in growth plates of chicks selected for a high incidence of TD. No differentiation-dependent expression of HIF-1alpha was observed in response to hypoxia, as demonstrated by the use of primary cultures of growth plate chondrocytes. In the normal growth plates, Hsp90 and Hsp70 were localized to the maturation zone. More cells expressed both Hsp in the TD lesion. In conclusion, we demonstrated that the TD growth plate, in contrast to the normal one, is hypoxic, probably because of the lack of vascularization. Hypoxia leads to an increase in the transcription factor HIF-1alpha, causing increases in the levels of Hsp90 and Hsp70.

Induction of tibial dyschondroplasia in turkeys by tetramethylthiuram disulfide (thiram).

Tibial dyschondroplasia (TD) is a prevalent skeletal abnormality associated with rapid growth rate in many avian species; it causes enormous economic losses and is an animal welfare problem. Tibial dyschondroplasia is characterized by the presence of a nonvascularized, nonmineralized lesion that extends from the epiphyseal growth plate into the metaphysis of the proximal tibiotarsal bones. The mechanisms underlying TD development are not known, although they have been extensively studied in broilers using different induction models. However, an effective model for TD induction in turkeys has never been described. The objective of this study was to establish such a model by using tetramethylthiuram disulfide (thiram), an agent that is frequently used in broilers to induce TD. We found that dramatically longer exposures to much higher concentrations of thiram were required to induce TD in turkeys vs. broilers. In contrast to broilers, in which 50 mg/kg of thiram induces a high incidence of severe TD within 10 d, in turkeys, an exposure to 400 mg/kg of thiram for 11 wk was necessary for the development of severe TD lesions. These results show different mechanisms for TD induction in these 2 closely related species, suggesting differences in TD etiology between them.

Use of a computer simulator for training children with disabilities in the operation of a powered wheelchair.

OBJECTIVE: The purpose of this study was to evaluate the ability of a basic driving simulator program to evaluate and train children with disabilities in their ability to operate a powered wheelchair. METHOD: With a rating scale of skills considered essential for safe and efficient wheelchair operation, 22 children 7 to 22 years of age with either progressive muscular dystrophy or cerebral palsy were evaluated in their ability to drive a powered wheelchair through a driving course. They were divided into two groups: one without prior experience driving a powered wheelchair and the other with experience. After the driving assessment with an actual powered wheelchair, the inexperienced drivers were trained on a joystick-controlled computer game in which they navigated through labyrinths similar in layout to their own school environment. A test maze was administered before and after this training. Both groups were then evaluated on their ability to drive a powered wheelchair through the driving course. RESULTS: The inexperienced drivers significantly increased their simulator scores over the training period. Their wheelchair driving performance was significantly better after the simulator training, although their performance remained poorer than that of the experienced drivers. CONCLUSION: A simulator program can assist in the development and evaluation of the skills required to operate a powered wheelchair.

Growth, digestibility, and enzyme activities in the pancreas and intestines of guinea-pigs fed on raw and heated soya-bean flour.

The nutritional effects of giving raw (RSF) or heated (HSF) soya-bean flour to young guinea-pigs were investigated in trials 1 and 2, in which the levels of dietary protein were 120 and 190 g/kg diet respectively. The growth rate of animals fed on RSF was lower than that of those fed on HSF. Growth retardation of guinea-pigs fed on RSF was accompanied by a lower apparent digestibility of the protein (0.49-0.53) compared with HSF (0.67-0.76) and lower food conversion efficiency. In RSF-fed animals, increasing dietary protein affected growth and food conversion efficiency negatively. The pancreas of animals fed on RSF and HSF was similar in weight but secreted less trypsin, chymotrypsin and amylase, in RFS-fed animals. It was concluded that the mechanism by which raw soya-bean negatively affects the growth rate of guinea-pigs by reducing the activity of intestinal enzymes, differs from that suggested for rats and chicks, but is similar to that of pigs and calves.

The failure of long-term feeding of raw soy flour, in the presence or absence of azaserine, to induce carcinogenic changes in the mouse pancreas.

The effects of feeding mice raw or heated soy flours or casein in the presence and absence of injected azaserine were investigated over a period of 18 months. Although the feeding of raw soy flour (compared with heated soy flour or casein) caused a significant inhibition of growth and an enlargement of the pancreas, there was no macroscopic evidence of pancreatic nodules in any of the six experimental groups. Microscopic examination of the pancreas revealed a somewhat higher (not significant) incidence of atypical acinar cell nodules in all animals injected with azaserine, but this difference was little influenced by the diets themselves. We concluded that raw soy flour itself has no carcinogenic effect on the mouse pancreas and does not enhance the sensitivity of the mouse pancreas to azaserine. Thus, it cannot be assumed that the appearance of pancreatic nodules constitutes an obligatory sequela of pancreatic hypertrophy and/or hyperplasia in all species of animals.

The effect of the long-term feeding of raw soy flour on the pancreas of the mouse and hamster.

The effects of the long-term feeding of mice and hamsters with raw (RSF) or heated (HSF) soy flours were investigated both in the presence and absence of chemical carcinogens. Mice which had been fed RSF for 18 months had enlarged pancreas but only a relatively low incidence of atypical acinar cell nodules (AACN). Mice on either RSF or HSF were also relatively resistant to the carcinogenic effects of azaserine which is known to produce a high incidence of AACN in rats. Hamsters which had been fed RSF for 15 months did not exhibit pancreatic enlargement and had a very low incidence of pancreatic tumors. Although the incidence of tumors in hamsters which had been injected with N-nitrosobis (2-oxopropyl)amine and maintained on HSF was very high (88%), those on RSF had a tumor incidence of less than 10%. Thus, there appears to be a marked difference in the response of the pancreas in different species of animals to the long-term effects of feeding RSF. This should be taken into consideration in evaluating the potential carcinogenicity of the trypsin inhibitors.

Effect of raw soy flour, feeding regime, and azaserine on rat pancreas.

The combined effects of feeding rats increasing amounts of raw soy four, feeding regime (ad libitum vs one meal per day), and injection of azaserine on the incidence of pancreatic nodules were investigated over a period of 12 months. Food consumption and body weights of meal-fed rats were lower than those of their ad libitum counterparts. The difference in body weight between the ad libitum and meal-fed rats became greater as the level of raw soy flour in the diet increased. Azaserine injections did not affect food consumption or body weight. The weights of the pancreas (gm/100 gm BW) increased parallel to the level of raw soy flour in the diet. The survival rate of rats on diets containing 19% and 42% raw soy flour was greater on the meal-fed regime than in the ad libitum group, but the converse was true when the diet contained 80% raw soy flour. The number and severity of pancreatic nodules observed in azaserine-injected animals surviving at the end of 12 months increased in relation to the level of raw soy flour in the diet and was enhanced at each level by meal feeding.

The effects of soy flour and N-nitrosobis (2-oxopropyl) amine on the pancreas of the hamster.

Syrian golden hamsters fed diets of raw or heated soy flour for 16 months had an incidence (4%) of pancreatic neoplasms in both groups. Animals injected with N-nitrosobis (2-oxopropyl) amine (BOP) and fed heated soy flour for 15 months developed a high incidence (88%) of microscopic benign and malignant neoplasms, primarily of ductal origin. This was in marked contrast to a similar group of BOP-injected animals which had been fed raw soy flour and in which the incidence of pancreatic neoplasms was less than 10%.

Availability of macro and microelements from edible domestic waste fed to sheep.

Young Awassi rams fitted with rumen cannula and T-shaped cannulas in duodenum and terminal ileum were used to study net absorption of calcium, phosphorus, magnesium, zinc, manganese, and copper from edible domestic waste in comparison with a conventional concentrate diet. Mean net absorptions (% of intake) of calcium, phosphorus, and magnesium from edible domestic waste and control diets were 15.3, -.9; 54.4, 37.0; and 46.4, 58.8. The small intestine was the major site for calcium and phosphorus absorption and the rumen for absorption of magnesium. Net absorption of zinc, manganese, and copper was low in both diets, probably as a result of excessive supply and low physiological demands. There was a net gain of manganese and copper and a net loss of zinc in the stomachs. In addition to potential for energy and protein, edible domestic waste supplies macro and microelements for ruminants, and thus, it can be regarded as a complete concentrate diet.

Growth, digestibility and enzymatic activities in the pancreas and intestines of hamsters fed raw and heated soy flour.

The nutritional effects of feeding raw and heated soy flour to young golden Syrian hamsters were investigated over a period of 32 days. Those animals fed raw soy flour grew much more poorly than those fed heated soy flour, an effect which was reflected in a lower food efficiency as well. Growth retardation of hamsters fed raw soy flour was accompanied by a lower apparent digestibility of the protein (54%) compared to heated soy flour (76%). The pancreases of animals fed raw soy flour were increased in size and had elevated levels of trypsin, chymotrypsin, amylase, and lipase. With the exception of trypsin activity in the small intestine, similar differences in enzyme activities between the raw and heated soy groups were generally found in the small intestine, cecum, large intestine, and feces. There was, however, a progressive decrease in these activities in the lower regions of the intestinal tract and feces. It is concluded that the hamster, in common with several other species of animals, is sensitive to the effects of the trypsin inhibitors in raw soy flour, and may provide a useful model for studying the long-term effects of trypsin inhibitors on the pancreas.

Effect of protein infused into sheep duodenum on activities of pancreatic proteases in intestinal digesta and on the absorption site of amino acids.

The effect of increasing the flow of protein amino acids to the small intestine on the activities of pancreatic proteases and on the absorption site of amino acids was examined in sheep equipped with intestinal cannulas at 0.05, 11 and 25 m from the pylorus. A single concentrated diet providing 85.1 g of protein amino acids/day was fed throughout the experiment. During period 1 this was the only source of protein. During period 2, a suspension of casein was infused into the duodenum through the 0.05-m cannula at a rate of 100 g/day, which supplied to the small intestine a total of 177 g amino acids/day. In period 1, 56.7 g of amino acids was absorbed per day along the entire small intestine, and 85% of it was absorbed in the anterior 11 m. Of the 143 g of amino acids absorbed per day during period 2, 63% was absorbed in the anterior 11 m. Ninety-four percent of the infused casein was absorbed. Increasing the flow of amino acids to the small intestine of sheep from 85 to 177 g/day resulted in an overall elevation in the activities of chymotrypsin, carboxypeptidase A and trypsin at the 11-m site but not the 25-m site. The greatest increase was obtained with chymotrypsin: 119 and 114% for the specific and total activities, respectively, and the least with the trypsin: 38 and 36% for the specific and total activities, respectively.