In vitro and in vivo effects of activated macrophage supernatant on distal limb wounds of ponies.

OBJECTIVE: To determine whether monokines produced by activated rabbit peritoneal macrophages can inhibit development of exuberant granulation tissue formation in distal limb wounds in ponies. DESIGN: Randomized block. ANIMALS: 5 castrated male ponies, 2 to 6 years old and weighing 140 to 190 kg. PROCEDURE: In vitro activity of cell-free rabbit peritoneal macrophage supernatant was determined after incubation of fibroblasts from the flank and the distal portion of limbs of horses and ponies. Tritiated thymidine was then added, and after reincubation, radioactivity was measured. After creation of a 4-cm2, full-thickness wound on the mid dorsal aspect of each metacarpus and metatarsus of each pony, in vivo activity of the macrophage supernatant was evaluated. Biopsy specimens were collected at random sites near a border of each wound at 4, 6, and 10 weeks after creation of the wounds. Treatment effects were evaluated on the basis of presence of exuberant granulation tissue requiring excision, number of times that excision was required, total area of the wound, epithelialized area, area of granulation bed, and histologic evaluation of the biopsy specimens. RESULTS: The macrophage supernatant effectively inhibited proliferation of equine fibroblasts in vitro. No significant in vivo treatment effects were found among the 4 treatment groups. CONCLUSION AND CLINICAL RELEVANCE: Monokines from stimulated rabbit peritoneal macrophages may have potential for improving wound healing in horses and ponies because of their effective inhibition in vitro of equine fibroblast proliferation.

Growth hormone (GH) receptor and GH-binding protein messenger ribonucleic acids with alternative 5'-untranslated regions are differentially expressed in mouse liver and placenta.

Two 5'-untranslated regions (5'UTRs) with distinctly different sequences, designated 5'UTR L1 and 5'UTR L2, were obtained by amplification of complementary DNA from mouse liver and placenta with primers complementary to sequences from the hormone-binding domain common to GH receptor (GHR) and GH-binding protein (GHBP) messenger RNAs (mRNAs). The presence of an open reading frame in the 5'UTR L2 and the high GC content of this sequence suggest that mRNAs containing this 5'UTR may be translated with a lower efficiency than those containing 5'UTR L1. Expression studies showed that 5'UTR L1 and 5'UTR L2 are present in GHR and GHBP mRNAs in both tissues. However, the relative expression of the two 5'UTRs differs between liver and placenta and in liver from different physiological states. The different expression patterns of the L1 and L2 5'UTRs predict that the corresponding 5'-noncoding exons of the GHR/GHBP gene are associated with different regulatory elements. The expression patterns of the 5'UTRs also indicate that there is a linkage between the 5'UTR present in GHR/GHBP gene transcripts and the alternative splicing of these transcripts to yield either GHR or GHBP mRNAs. The 5'-noncoding exon used for transcription of the GHR/GHBP gene, therefore, may be involved in regulating both the ratio of GHR to GHBP transcripts and the efficiency of translation of these transcripts. Transcription from the different 5'-noncoding exons of the GHR/GHBP gene thus may be a critical element in the regulation of the expression of GHR and GHBP and thereby in the control of the responses of different tissues to GH.

A hypoglycemic factor from the corpus cardiacum of the American cockroach Periplaneta americana.

Thin-layer chromatography of cockroach corpora cardiaca (cc) extracts revealed a fraction (F1 in solvent system 1) which had hyperlipemic activity in locusts and which differed in mobility from synthetic locust adipokinetic hormone I (AKH I). Fractions 4 and 5 exhibited hypolipemic activity when injected into locusts. Pooled samples of these fractions caused a diminution of carbohydrate levels in the cockroach. Bovine insulin mimicked both these effects. No change in hemolymph lipid was evident in the cockroach after injection of either CC fractions 4 and 5 or bovine insulin. Both the active fraction from the cockroach CC and bovine insulin caused a decrease in hemolymph carbohydrate in neck-ligated locusts. Injection of locust hypolipemic hormone, separated from locust CC storage lobes by TLC, into neck-ligated locusts also caused a decrease in hemolymph carbohydrate concentration. Normal locusts showed no change in hemolymph carbohydrate.

Cytomorphological lesions induced by chemotherapeutic agents in transitional cell carcinoma in vitro.

Transitional cell carcinoma from 20 patients and two human cell lines were maintained in short-term tissue culture. Each was studied ultrastructurally before and after incubation with cisplatinum, adriamycin, or mitomycin C. Sequential ultrastructural changes were noted and were found to be specific for each agent tested. Ultrastructural changes in the nucleoli were produced by exposure to cisplatinum or mitomycin C; alterations in the heterochromatin of the nuclei were characteristic of treatment with adriamycin. The changes in the nucleoli seen with cisplatinum have not been described previously and support an alkylating property as a mechanism of action. Intravesical chemotherapeutic agents are now commonly used in clinical treatments. The morphological changes produced by these agents are specific and may be seen in the clinical setting.

Transitional explant reduction assay--a new in vitro testing system for intravesical chemotherapy.

Eighty-five tumors from 49 patients with transitional cell carcinoma were examined in a model for in vitro sensitivity using tumor explant reduction assay. Individual sensitivity patterns were obtained for each tumor tested. Explants were most frequently found to be sensitive to cis-platinum (49 per cent) and least frequently sensitive to thiotepa (20 per cent). Thirteen patients were studied on multiple sequential intervals, with resistance to specific agents noted to develop in 53 per cent of the patients. To date, a prospective cross-over study has been conducted on seven patients with 15 correlates. Overall, clinical correlation of this assay for both resistance and sensitivity is 93 per cent. We report a new assay for transitional cell carcinoma of the bladder that is adaptable to small specimen samples and provides adequate material for testing 73 per cent of the time.

Stimulus selection in passive avoidance learning and retention: weanling, periadolescent, and young adult rats.

Periadolescent rats exhibit a number of behavioral differences in comparison with younger or older animals. For instance, periadolescents tend to show enhanced acquisition of simple active avoidance tasks, but impaired acquisition of more complex appetitive and aversive discriminations. In this experiment, rats were trained on a simple passive avoidance task at one of three ages, as weanlings (25 days), periadolescents (35 days), or young adults (45 days). Training occurred in the presence of both a redundant discriminative stimulus and a specified, redundant contextual stimulus. The periadolescents did not differ from either younger or older rats in rate of learning the passive avoidance task. The retention performance of these animals was then tested following a change in either, neither, or both of the redundant cues. When a measure of performance that controls for baseline activity was used, it was observed that periadolescents were not disrupted by a change in the redundant discriminative stimulus, a cue change that clearly disrupted performance in 25- and 45-day-old animals, and tended to be more disrupted by the contextual change than younger or older rats. It is hypothesized that the alterations in performance exhibited by periadolescents may be related to an ontogenetic alteration in stimulus selection modulated by the catecholaminergic systems.

Wall climbing, odors from the home nest and catecholaminergic activity in rat pups.

The present investigation examined the relationship among wall climbing, home nest odors, circadian rhythmicity and catecholaminergic activity. In the first experiment, a catecholaminergic agonist, amphetamine, was observed to induce wall climbing in 10- but not 15-day old rat pups. It was observed in the second experiment that an alpha-adrenergic antagonist, phentolamine, attenuated wall climbing elicited by foot shock in 10-day old rat pups without affecting general locomotor activity. However, haloperidol, a dopaminergic antagonist, was observed to decrease both shock-elicited wall climbing and general locomotor activity in rat pups at this age. These results support the hypothesis that wall climbing may be partially mediated by catecholaminergic activity. Home nest odors decreased both amphetamine-induced wall climbing for animals tested during the dark cycle in Experiment 1 and shock-induced wall climbing for subjects tested both during the light and dark phases in Experiment 2. This attenuation by home nest odors of both drug- and shock-elicited wall climbing suggests the further hypothesis that home nest odors may influence the level of catecholaminergic activity.

Lymphocyte-tumor cell interaction in patients with head and neck cancer.

Peripheral lymphocytes from 12 patients with squamous cell carcinoma of the lead and neck were incubated with autologous tumor explants. Four of the 12 patients demonstrated lymphocyte induced tumor cytotoxicity. These lymphocytes adhered to the tumor cells and deposited a radioactive label from their surface onto tumor cells. The deposition of this label was associated with tumor death. Tissue sections from those patients who demonstrated lymphocyte cytotoxicity showed a marked plasmacytic infiltration. This was in contrast to non-responders where only a desmoplastic tissue response was observed with few inflammatory cells.

Ultrastructure of lymphocyte tumor cell interaction with localization of cell-bound antibody by ferritin labeling.

Splenic lymphocytes derived from Walker carcinoma-bearing rats were harvested and incubated with Walker carcinoma cells growing in tissue culture. The sequence of events leading to target cell death was studied by phase microscopy and scanning and transmission electron microscopy. The sensitized lymphocytes adhere to the tumor cells by multiple cytoplasmic appendages, but no ultrastructural changes are seen at this interface. After 1 hr these lymphocytes release cytoplasmic components consisting of membrane-lined vesicles, cell membranes, endoplasmic reticulum, and cytoplasmic material. This material adheres closely to the surface of the tumor cells and is subsequently seen within the cytoplasm of the tumor cell. The tumor cells then undergo degenerative changes and cell death occurs in 24 to 36 hr. The lymphocyte-derived material appears to contain immunoglobulin components as determined by specific ferritin labeling.

Capillary blood collection in haematology.

A capillary blood collection technique which facilitates the estimation of routine haematological parameters, including platelet count and sedimentation rate, is described. The technique requires 0.5 ml of blood, allows closer reproducibility than pipette collection methods, is suitable for monitoring blood counts in patients receiving cytotoxic agents, and can be integrated with semi-and fully-automated production lines.