John W. Madden

Administration of beta-aminopropionitrile to human beings with urethral strictures: A preliminary report

(1) No toxic signs or symptoms and no unusual laboratory determinations were observed in five patients with posterior urethral strictures treated by dilatation of the urethra and administration of 1 gm per day of beta-aminopropionitrile (BAPN) for twenty-one days. (2) Patients treated with 1 gm daily of BAPN for twenty-one days showed an increase in cold saline-extractable and acid-extractable collagen in proplast sponge-collected and dermal scar tissue comparable with that reported previously after doses of 3 to 5 gm of BAPN daily. (3) Significant reduction in the breaking strength of newly synthesized connective tissue was observed in patients treated with BAPN. (4) None of the patients in this study showed abnormalities in net collagen synthesis or in synthesis of noncollagenous protein. (5) The difference in the results of this study and two previous trials of BAPN in human beings which were discontinued because of toxic and or hypersensitivity signs and symptoms is hypothesized to be the result of development of highly purified BAPN fumarate.

SOFT TISSUE RESPONSE TO PROPLAST: QUANTITATION OF SCAR INGROWTH

We compared, histologically and biochemically, the effects of blood pretreatment vs. no pretreatment on the ingrowth of scar into subcutaneously implanted blocks of Proplast in rats. No significant differences were found.

Activity and Extractability of Lysyl Oxidase and Collagen Proteins in Developing Granuloma Tissue

Summary The total activity of lysyl oxi-dase and its extractability into phosphate and 4 M and 8 M urea solutions were studied in granuloma tissue formed around sub-cutaneously implanted polyvinylalcohol sponges in rats and harvested on days 10, 18, 26, and 34. Furthermore, total collagen content and fractions of collagen extractable into 0.45 M NaCl (NSC) and 0.5 M acetic acid (ASC) were studied. Lysyl oxidase activity was assayed by measuring the release of tritium from a substrate biosynthetically labeled with H3-6-Lysine. Biochemical data were compared to the histology of the tissue. With the maturation of the granuloma tissue, the total activity of lysyl oxidase significantly decreases, and its extractability into phosphate significantly increases. Extractability into 4M urea was the highest in 10-day-old granuloma and the lowest in 34-day-old tissue. Only 80% of the total activity was extracted into 4 M urea medium. Total collagen content and ASC fraction increased continuously, and NSC decreased from 2.5% to 1.2% in 34-day-old tissue. We conclude that the highest activity of lysyl oxidase in young granuloma is related to a high proportion of fibroblasts and that the enzyme at this time is tightly bound to the pellet, possibly to the collagen substrate.

Toxicity and metabolic effects of 3,4-dehydroproline in mice.

Abstract The acute and chronic effects of 3,4,-dehydroproline (DHP) were investigated in mice. Given as a single dose ip, DHP is 13 times more toxic than 1-proline, the LD50 being 0.62 ± 0.05 g/kg. Ultrastructural changes observed in liver hepatocytes and granuloma tissue fibroblasts at various time intervals after single injections of DHP demonstrated cytotoxic effects involving smooth endoplasmic reticulum, mitochondria, and rough endoplasmic reticulum. Chronic effects of DHP (12 mg/100 g/12 hr, ip) on collagenous and noncollagenous protein metabolism were studied in liver, normal skin, dermal wounds, and polyvinyl alcohol-induced granuloma tissue. To amplify the effect of DHP, all animals were kept on proline-deficient diets. This diet, however, did not affect the size of the free proline pool in the liver. Chronic DHP administration reduced the quantity and rate of synthesis of noncollagenous protein in granuloma tissue and inhibited noncollagenous protein synthesis in skin. DHP had no measurable effect on liver mitochondrial or microsomal protein metabolism. Although the breaking strength of skin wounds was significantly higher after DHP, no effect on collagen synthesis or accumulation was observed in any system. Complete amino acid analysis of purified collagen from skin showed negligible incorporation of DHP. We conclude that chronic in vivo administration of DHP is toxic and does not interfere with collagen accumulation in normal or repaired tissues.

Soft tissue response to blood-impregnated Proplast.

The effects of Proplast implants on the soft tissues of rats were studied. The effects from Proplast implants which had been impregnated with blood, plasma, or saline were compared to the effects from implants which had received no pretreatment. No significant differences were found.

Effect of cis-hydroxyproline on collagen and other proteins in skin wounds, granuloma tissue, and liver of mice and rats

Abstract Acute and chronic pharmacologic effects of cis-hydroxyproline (cis-Hyp) were studied in mice and rats. Given as a single dose intraperitoneally, cis-Hyp is relatively non-toxic and the LD50 approaches that of l-proline. Chronic administration of cis-Hyp (200 mg/kg/day) to mice did not affect breaking strength of dermal wounds or content and specific activity of noncollagenous protein in the liver. Although cis-Hyp produced a significant decrease tn noncollagenous protein content of polyvinyl alcohol sponge induced granuloma tissue, collagen content and rate of synthesis in granuloma tissue and liver were increased significantly. Rats given cis-Hyp (200 mg/kg/day subcutaneously in divided doses every 12 hr) and control animals demonstrated identical hepatic microsomal protein, cytochrome P450 and cytochrome b5 content. Breaking strength of dermal wounds was increased significantly in treated animals. Ultrastructural analysis of rat liver failed to demonstrate any structural alterations of cellular organelles due to cis-hyp treatment. We conclude that chronic in vivo administration of cis-Hyp does not inhibit collagen synthesis or accumulation in normal or repaired tissues and does not decrease the breaking strength of skin wounds. Other effects on protein metabolism could result from nonspecific, chronic toxicity of cis-Hyp.

The Effect of Multiple Lathyrogenic Agents Upon Wound Healing in Rats

Summary 1. Simultaneous administration of BAPN and d-penicillamine in low nontoxic doses produced a significant additive inhibition of cross-bonding in collagen of healing wounds. Additive effect was not observed following administration of high doses of β-aminopropionitrile and d-penicillamine. 2. Toxic signs in rats, including partial alopecia, diarrhea, failure to gain weight, began to appear when the dose of BAPN exceeded 40 mg/100 g body weight/day. 3. Toxicity in rats treated with d-penicillamine was limited to weight loss and was noted when doses greater than 150 mg of d-penicillamine/100 g of body weight/day were administered. 4. These results support the hypothesis that d-penicillamine and BAPN act at different stages in the maturation of collagen and suggest that simultaneous administration of low nontoxic doses of several lathyrogenic agents may be more useful in treating human beings than large doses of a single agent.

The management of acute thermal injuries to the upper extremity

In summary, although thermal injuries of the upper extremity can produce permanent functional compromise, intelligent management of burns of the hands and fingers can produce fully functional and cosmetically acceptable hands. When the dorsal aspects of the hands alone are injured by thermal insult, the immediate excision of the burned tissue and replacement by split-thickness skin grafts will produce functional and esthetically satisfying results. Should immediate excision and grafting be contraindicated because of massive burns of the body, management by application of topical antibacterial agents and dressings is possible. With this treatment, carefully supervised exercises are mandatory. Revisions of hypertrophic and contracted scars can be done at a later date. The stiff and painful hand is not the direct consequence of thermally destroyed skin, but is due instead to the metabolic and biologic complications of unhealed burn wounds. It follows, therefore, that the burn illness should preferably be terminated by prompt excision of the eschar and wound closure by the application of split-thickness skin grafts.

Effect of 1,10-phenanthroline and desferrioxamine in vivo on prolyl hydroxylase and hydroxylation of collagen in various tissues of rats☆

Abstract We studied prolyl hydroxylase (PH) activity in various tissues of rats after systemic and local administrations of 1,10-phenanthroline and Desferrioxamine. We assayed PH in tissue extracts without adding iron to the incubation medium; we ascertained hydroxylation of collagen by measuring 3 H release from biosynthetically 3,4- 3 H-proline-labeled collagen substrate. We studied the effect of both Fe 2+ and Fe 3+ chelating agents in tissues with a relatively high rate of collagen synthesis such as fetal skin and skin of newborn rats, 17-β-estradiol-stimulated uterus of immature rats, and carrageenan granuloma tissue. In addition, in some organs we studied collagen hydroxylation by measuring the ratio of proline to hydroxyproline either in highly purified samples or in the digest from samples treated with protease-free collagenase. Finally, the extent of underhydroxylation of collagen from some tissues of rats treated with either one or both chelating agents was determined using a 3,4,- 3 H-proline-labeled collagen as a substrate to partially purified PH. Neither systemic nor local injections of 1,10-phenanthroline or Desferrioxamine alone inhibited PH in any of the models. But simultaneous administration of both agents inhibited PH and hydroxylation of collagen in some models. Although local injections of 1,10-phenanthroline into granuloma tissue did not inhibit PH activity, hydroxylation of collagen synthesized in the granuloma from 4 to 16 hr after injection was reduced significantly. This seeming discrepancy is explained by the finding that PH is active even without the addition of Fe 2+ in the assay medium; a strong reducing environment (α-ketoglutarate, ascorbic acid) reduces Fe 3+ , thus providing ferrous ions essential for PH activity. Both forms of iron, therefore, must be chelated simultaneously to affect PH activity in the assay system. In some experiments, within 8 hr after systemic administration of 1,10-phenanthroline, the activity of PH in skin almost doubled and remained significantly elevated for 40 hr. The mechanism responsible for this phenomenon is not known. The results of this study indicate that previously reported inhibitory effects of some Fe 2+ chelating agents on collagen synthesis in various models of fibrosis may not be related to inhibition of PH or to collagen hydroxylation.

Morphologic aspects of experimental esophageal lye strictures. I. Pathogenesis and pathophysiologic correlations.

Abstract Esophageal lye burns in dogs alter the esophageal wall in a predictable way. Tight morphologic stricture may develop in a few weeks, but only if the initial injury destroys the full thickness of the muscularis. Stricture is the result of deformity of the whole healing wall rather than massive proliferation of scar tissue, is not noticeably related to persistent mucosal ulceration, and greatly interferes with adequate food intake. However, functional and radiographic abnormalities may be quite severe even when the esophageal lumen is minimally narrowed on quantitative morphologic examination; factors such as edema and peristaltic dysfunction may account for disparities between structural and functional effects of lye injury.