An inbred line of transgenic mice expressing an internally deleted gene for type II procollagen (COL2A1). Young mice have a variable phenotype of a chondrodysplasia and older mice have osteoarthritic changes in joints.

Studies were carried out on a line of transgenic mice that expressed an internally deleted COL2A1 gene and developed a phenotype resembling human chondrodysplasias (Vandenberg et al. 1991. Proc. Natl. Acad. Sci. USA. 88:7640-7644. Marked differences in phenotype were observed with propagation of the mutated gene in an inbred strain of mice in that approximately 15% of the transgenic mice had a cleft palate and a lethal phenotype, whereas the remaining mice were difficult to distinguish from normal littermates. 1-d- and 3-mo-old transgenic mice that were viable showed microscopic signs of chondrodysplasia with reduced amounts of collagen fibrils in the cartilage matrix, dilatation of the rough surfaced endoplasmic reticulum in the chondrocytes, and decrease of optical path difference in polarized light microscopy. The transgenic mice also showed signs of disturbed growth as evidenced by lower body weight, lower length and weight of the femur, decreased bone collagen, decreased bone mineral, and decreased resistance of bone to breakage. Comparisons of mice ranging in age from 1 d to 15 mo demonstrated that there was decreasing evidence of a chondrodysplasia as the mice grew older. Instead, the most striking feature in the 15-mo-old mice were degenerative changes of articular cartilage similar to osteoarthritis.

Alterations in prodynorphin gene expression and dynorphin levels in different brain regions after chronic administration of 14-methoxymetopon and oxycodone-6-oxime.

Previous studies showed that opioid drugs-oxycodone-6-oxime and 14-methoxy-5-methyl-dihydromorphinone (14-methoxymetopon)-produced less respiratory depressive effect and slower rate of tolerance and dependence, respectively. It was also reported that morphine decreased the prodynorphin gene expression in the rat hippocampus, striatum and hypothalamus. In this study, we determined the prodynorphin gene expression and dynorphin levels in selected brain regions of opioid tolerant rats. We found that in the striatum morphine decreased, while oxycodone-6-oxime increased and 14-methoxymetopon did not alter the prodynorphin gene expression. In the nucleus accumbens, morphine and oxycodone-6-oxime did not change, while 14-methoxymetopon increased the prodynorphin gene expression. In the hippocampus both oxycodone-6-oxime and 14-methoxymetopon enhanced, whereas morphine did not alter the prodynorphin gene expression. In the rat striatum only oxycodone-6-oxime increased dynorphin levels significantly in accordance with the prodynorphin mRNA changes. In the hippocampus both opioid agonists increased the dynorphin levels significantly similarly to the augmented prodynorphin gene expression. In ventral tegmental area only 14-methoxymetopon increased dynorphin levels significantly. In nucleus accumbens and the temporal-parietal cortex the changes in the prodynorphin gene expression and the dynorphin levels did not correlate. Since the endogenous prodynorphin system may play a modulatory role in the development of opioid tolerance, the elevated supraspinal dynorphin levels appear to be partly responsible for the reduced degree of tolerance induced by the investigated opioids.

Expression and prognostic value of alpha-, beta-, and gamma-catenins indifferentiated thyroid carcinoma.

Catenins (alpha, ss, and gamma) are a group of intracellular cell adhesion molecules that unite cytoskeleton with extracellular adhesion system. Abnormal expression of these molecules may have prognostic relevance in various carcinomas, including differentiated thyroid carcinoma (DTC). We have, therefore, evaluated the prognostic value of alpha-, ss-, and gamma-catenins along with traditional risk factors in 206 consecutive DTC patients by immunohistochemistry. Papillary carcinomas showed normal staining pattern for alpha-, ss-, and gamma-catenins in 124 (60%), 136 (67%), and 94 (46%) cases, respectively. Follicular carcinomas expressed alpha-, ss-, and gamma-catenins normally in 16 (48%), 18 (55%), and 8 (32%) cases, respectively. Follicular type of tumor showed more often reduced staining for all catenins than papillary carcinoma (P: = 0.009, P: = 0.004, and P: = 0.002, respectively). Age (>60 yr) and pTNM-stage were related to reduced alpha- and ss-catenin expression levels (P: = 0.027 and P: = 0.026, respectively) and larger size of the tumor to reduced ss- and gamma-catenin expressions (P: = 0.039 and P: = 0.007, respectively). Nodal metastases at the time of primary treatment related to reduced alpha-catenin expression and distal metastases to reduced ss- and gamma-catenin staining signals (P: = 0.022, P: = 0.014, and P: = 0.039, respectively). Reduced alpha-catenin associated with tumor recurrence (P: = 0.002) and reduced ss-catenin with cancer-related mortality (P: = 0.005). The multivariate analysis for recurrence-free survival showed that alpha-catenin and serum thyroglobulin level 1 yr after primary treatment were prognostic of recurrent disease (hazards ratio, 3.42, P: = 0.022; and hazards ratio, 10.03, P: = 0.0001). In addition, alpha-catenin retained its prognostic significance in low-stage patients (P: = 0.0151). We propose that the evaluation of alpha-catenin expression by immunohistochemistry in DTC patients has prognostic value in addition to that obtained by traditional prognostic factors.

A new method for obtaining viable cells from dermal infiltrates. A study on 2,4 dinitrochlorobenzene induced contact dermatitis.

Experimental contact dermatitis has been induced in 2,4 dinitrochlorobenzene (DNCB) sensitized guinea pigs. The developing dermal infiltrate was excised and the infiltrating cells were obtained by mechanical extraction alone as well as by the combination with collagenase and elastase treatment. The most viable cells appeared in the elastase and mechanically extracted samples and the least in those subjected to mechanical treatment alone. The most cells in the enzyme-treated samples were present 24 h after re-exposure of the sensitized animals to DNCB consisting mainly of lymphocytes and of polymorphonuclear granulocytes. The optimum conditions for the action of enzymes including optimum duration of the treatment, buffer milieu, aspecific proteolytic effect on foreign substrate and action on T and B cell receptors have been elaborated. It was concluded that 80 min of collagenase treatment with gentle mechanical extraction under specified conditions does not affect any measurable immunologic properties of the liberated cells resulting in the second best yield. A comparison of these data with earlier reports and their significance is being discussed.

The progression of burn depth in experimental burns: a histological and methodological study.

UNLABELLED: This study was designed to create a reproducible model for experimental burn wound research in pigs. Previously, the thicker paraspinal skin has been used. We used the more human-like ventral skin to create burns of different depths. Contact burns were created to 11 pigs using a brass plate heated to 100 degrees C in boiling water. Different contact times were used to create burns of different depths. In pigs 1-6, the follow-up time was 72 h and in pigs 7-11 24 h. Burn depth was determined by histology. Histologically, samples were classified into five anatomical layers: epidermis, upper one-third of the dermis, middle third of the dermis, deepest third of the dermis and subcutaneous fat. The location of both thromboses and burn marks were evaluated, respectively. The 1 s contact time lead to a superficial thermal injury, 3 s to a partial thickness and 9 s to a full thickness injury. A progression of burn depth was found until 48 h post-injury. The intra-observer correlation after repeated histological analyses of burn depths by the same histopathologist and the repeatability of burn depth creation yielded kappa coefficients 0.83 and 0.92, respectively. CONCLUSION: a reproducible burn model for further research purposes was obtained.

Effects of intra-articular injections of bufexamac suspension in healthy horses.

OBJECTIVE: To evaluate the effects of intra-articular (IA) injections of bufexamac in horses, focusing particularly on the effects of bufexamac on articular cartilage. ANIMALS: 20 Standardbreds. PROCEDURE: Horses were randomly allocated into 4 groups consisting of 5 horses each, and 20, 60, or 100 mg of bufexamac or 1 ml of sterile saline (0.9% NaCl) solution (control) was injected into 1 intercarpal joint at weekly intervals for 6 treatments (days 0, 7, 14, 21, 28, and 35). Clinical signs and results of hematologic, serum biochemical, and synovial fluid (SF) analyses and radiography were used to evaluate treatment effects. On day 49, all horses were euthanatized; gross necropsy and histologic examinations of internal organs and articular tissues were performed. Glycosaminoglycan concentration of the articular cartilage was evaluated in safranin O-stained sections by use of a semiquantitative microspectrophotometric method. RESULTS: No systemic signs were observed. Temporary mild to moderate heat and effusion were the only clinical signs observed in a number of joints after IA injections and more often only in the 100 mg group, compared with controls. The 100 mg dose resulted in significant increases in SF WBC counts, with relative neutrophilia and SF total protein concentration 24 hours after injection (day 1). No lesions suggestive of toxic effects were detected at necropsy or on histologic examination. No changes in articular cartilage glycosaminoglycan concentration were detected. CONCLUSIONS AND CLINICAL RELEVANCE: Six injections of 20, 60, or 100 mg of bufexamac at weekly intervals did not cause any untoward systemic or local effects. These data suggest that bufexamac is a safe nonsteroidal anti-inflammatory drug for IA administration in horses.

Spinal interaction between the highly selective µ agonist DAMGO and several δ opioid receptor ligands in naive and morphine-tolerant mice.

Since the discovery of opioid receptor dimers their possible roles in opioid actions were intensively investigated. Here we suggest a mechanism that may involve the µ-δ opioid heterodimers. The exact role of δ opioid receptors in antinociception and in the development of opioid tolerance is still unclear. While receptor up-regulation can be observed during the development of opioid tolerance no µ receptor down-regulation could be detected within five days. In our present work we investigated how the selective δ opioid receptor agonists and antagonists influence the antinociceptive effect of the selective µ receptor agonist DAMGO in naïve and morphine-tolerant mice. We treated male NMRI mice with 200 µmol/kg subcutaneous (s.c.) morphine twice daily for three days. On the fourth day we measured the antinociceptive effect of DAMGO alone and combined with delta ligands: DPDPE, deltorphin II (agonists), TIPP and TICPψ (antagonists), respectively, administered intrathecally (i.t.) in mouse tail-flick test. In naive control mice none of the δ ligands caused significant changes in the antinociceptive action of DAMGO. The treatment with s.c. morphine resulted in approximately four-fold tolerance to i.t. DAMGO, i.e. the ED50 value of DAMGO was four times as high as in naive mice. 500 and 1000 pmol/mouse of the δ1 selective agonist DPDPE enhanced the tolerance to DAMGO while 1000 pmol/mouse of the δ2 selective agonist deltorphin II did not influence the degree of tolerance. However, both δ antagonists TIPP and TICPψ potentiated the antinociceptive effect of i.t. DAMGO, thus they restored the potency of DAMGO to the control level. The inhibitory action of DPDPE against the antinociceptive effect of DAMGO could be antagonized by TIPP and TICPψ. We hypothesize that during the development of morphine tolerance the formation of µÎ´ heterodimers may contribute to the spinal opioid tolerance. δ ligands may affect the dimer formation differently. Those, like DPDPE may facilitate the dimer formation hence inhibit the antinociceptive effect of DAMGO by causing virtual µ receptor down-regulation. Ligands that do not affect the dimer formation do not influence antinociception either but ligands with the presumed capability of disconnecting the dimers may decrease the spinal tolerance to DAMGO.