N-acetylcysteine in pig liver transplantation from non-heart-beating donors.

Lipid peroxidation due to oxygen free radicals (OFR) seems to play a major role in loss of liver graft viability after warm ischemia, preservation, and transplantation. N-acetylcysteine (NAC) is an antioxidant that has a direct effect on OFR, and is also a glutathione precursor, another antioxidant. This study was designed to evaluate the efficacy of NAC in preventing ischemia-reperfusion damage of liver grafts harvested from non-heart-beating donors. Liver transplantation was performed on pigs divided into five groups: group 1 (control group; n=5) received livers from heart-beating donors; livers were subjected to 30 min of warm ischemia in groups 2 (n=3, no NAC) and group 3 (n=3; NAC treatment); warm ischemia time lasted 60 min in groups 4 (n=4; no NAC) and 5 (n=5; NAC treatment). Studied parameters included graft survival for more than 3 days, aspartate aminotransferase plasma levels, liver histology, and hepatic total glutathione concentrations. Graft survival was 100% in groups 1, 2, and 3, 0% in group 4, and 20% in group 5. NAC treatment did not influence initial mean aspartate aminotransferase release which was greater in warm ischemic livers than in controls. NAC treatment had no effect on liver hepatic total glutathione after reperfusion of animals receiving warm ischemic grants. Finally, no effect on liver histology was observed with NAC treatment. Our study suggests that in liver transplantation from non-heart-beating donors, NAC has no effect in both graft viability and lipid peroxidation. The role of OFR in primary dysfunction of transplanted warm ischemic livers remains controversial.

A novel mechanism for isolated central hypothyroidism: inactivating mutations in the thyrotropin-releasing hormone receptor gene.

Isolated central hypothyroidism, characterized by insufficient TSH secretion resulting in low levels of thyroid hormones, is a rare disorder. We report a boy in whom isolated central hypothyroidism was diagnosed at 9 yr of age. Complete absence of TSH and PRL responses to TRH led us to speculate that he had an inactivating mutation of the TRH receptor gene. The patients' genomic DNA was isolated, and the entire coding region of the TRH receptor was amplified by the PCR and sequenced directly. Confirmation of the mutations and haplotyping of the family was performed using restriction enzymes. The biological activity of the wild-type and mutated TRH receptors was verified by evaluating the binding of labeled TRH and stimulation by TRH of total inositol phosphate accumulation in transfected HEK-293 and COS-1 cells. The patient was found to be a compound heterozygote, having inherited a different mutated allele from each of the parents; both mutations were in the 5'-part of the gene. Mutated receptors were unable to bind TRH and to activate total inositol phosphate accumulation. Our report is the first description of naturally occurring inactivating mutations of a G protein-coupled receptor linked to the phospholipase C second messenger pathway. The prevalence and phenotypic spectrum of TRH receptor mutations in isolated central hypothyroidism remain to be established.

Identification of human growth hormone-releasing hormone receptor splicing variants.

The expression pattern of the receptor for human GHRH (hGHRHr) was investigated in normal human tissues and in pituitary and ovarian tumors by reverse transcription-polymerase chain reaction. Three transcriptional variant forms of the hGHRHr were found to be expressed in the normal pituitary and in GH-secreting pituitary tumors. Form a is identical to the previously reported hGHRHr, whereas forms b and c are not described in the literature. Form b and c are predicted to be transcribed into two truncated proteins. We amplified genomic DNA using primers designed from the complementary DNA of hGHRHr. A 2-kilobase genomic DNA fragment was cloned that contains part of the hGHRHr gene consisting of two exons, e864 and e934, separated by three introns, i863, i933, and i1025. Alternative splicing of i1025 was responsible for three variant forms of hGHRHr. Nonsecreting pituitary tumors showed an abnormal expression of the hGHRHr, probably due to alternative usage of exons at the 5'-end of the gene, although they also expressed the three variant forms. No hGHRHr expression was identified in a human mammosomatotroph cell line insensitive to GHRH, in normal human liver or ovary, or in various human ovarian tumors.

Mammosomatotroph adenoma causing gigantism in an 8-year old boy: a possible pathogenetic mechanism.

The pathophysiology of mammosomatotroph adenomas remains unclear. We studied a mammosomatotroph adenoma removed from an 8-year old boy with a 5-year history of growth acceleration and acromegalic gigantism at presentation. Elevated basal GH (mean 28 micrograms/l) and PRL (mean 120 micrograms/l) plasma levels were observed, as well as paradoxical responses of GH to L-dopa, TRH and oral glucose administration; PRL was reduced by L-dopa and slightly increased by TRH; GHRH stimulated release of both GH and PRL. Two operations were required to remove the very large tumour and the patient was treated with bromocriptine before the second. Hormonal secretion by tumour explants in culture was evaluated under basal conditions and after stimulation or inhibition. High levels of GH and PRL were secreted for up to 24 days. Furthermore, GHRH and TRH caused a dose-related stimulation of both hormones, while somatostatin and dopamine were effective in suppressing either basal or stimulated hormone release only at very high (microM) concentrations. Intracellular events were studied by determination of the guanosine triphosphate binding (G) protein levels and adenylate cyclase (AC) activity in the tumour tissue. Before bromocriptine treatment, AC activity was very high in the tumour and could be further stimulated by various agents; very high levels of the AC-stimulatory G protein alpha subunit Gs alpha and very low amounts of the AC-inhibiting G protein alpha subunit Gi3 alpha and of the phospholipase C-stimulating G protein alpha subunit Gq alpha were found in the tumour. After bromocriptine, baseline AC activity was normalized and could no longer be stimulated; Gs alpha and Gi3 alpha levels were unchanged while those of Gq alpha were normalized. Screening of tumour DNA after amplification by polymerase chain reaction followed by single-strand conformational polymorphism analysis did not reveal any mutations in the hot spots of G protein alpha subunits (alpha s, alpha i2, alpha o2 and alpha 11) genes or in the H-ras and p53 genes. Gs alpha and GH transcription factor-1 (pit-1) expression were evaluated by amplification of cDNA. While the mRNA expression of pit-1 decreased after bromocriptine treatment, that of Gs alpha increased. These data suggest the possibility of an oncogenic process involving overexpression of Gs alpha, resulting in chronic activation of adenylate cyclase. Furthermore, our results suggest that the anti-secretory and anti-proliferative effects of bromocriptine may be mediated through a decrease in Pit-1 secondary to the inhibition of adenylate cyclase activity.

Cellular distribution of G protein Go alpha in pituitary lactotrophs: effects of dopamine.

Membrane-bound GTP-binding (G) proteins mediate signal transduction in a variety of cell systems. The exact mechanisms of G proteins action are still under investigation but they appear to involve effectors located in the plasma membrane as well as in other parts of the cell. With this study, we investigated the cellular and ultrastructural localization of G protein subunits, and particularly of Go alpha, in normal rat anterior pituitaries and in estrone-induced rat adenomatous lactotrophs. We also evaluated the effects of Go alpha cellular redistribution in rat adenomatous lactotrophs following short-term exposure to dopamine (DA). Using the Protein A-gold (PAG) methodology, Go alpha was found to be present in the cysternae of the endoplasmic reticulum of normal pituitary cells and of adenomatous lactotrophs. In the latter, Go alpha could be co-localized with prolactin (PRL). By immunoblots, using specific antisera, significant amounts of Go alpha and Gs42 alpha, together with smaller amounts of Gi alpha, Gs47 alpha and G beta were found to be present in the uncontaminated supernatant fraction of adenomatous lactotrophs. Unexpectedly, exposure of the cells to DA induced a rapid and short-lived decrease in the cytosolic fraction of Go alpha and G beta associated with a decrease of PRL release. Since cytosolic Go alpha can be ADP-ribosylated by pertussis toxin (PT) and is therefore in a heterotrimeric form, our data suggest that the soluble Go protein may play a role during lactotrophs' exposure to an inhibitor of PRL release, perhaps through its relocalization after being internalized with the D2 receptor or by being used for interaction with intracellular and/or membrane-bound effectors.

MMQ cells: a model for evaluating the role of G proteins in the modulation of prolactin release.

It is well known that dopamine (DA) inhibits while vasoactive intestinal peptide (VIP) and angiotensin II (ANG II) stimulate prolactin (PRL) release from normal anterior pituitary lactotrophs; however, elucidation of the intracellular mechanisms involved in these effects has been hindered by the cellular heterogeneity of the anterior pituitary. MMQ cells, isolated from the PRL-secreting rat pituitary tumor 7315a is an interesting model since they only secrete PRL. In order to determine whether and which GTP-binding (G) proteins are involved in the modulation of cyclic 3',5'-adenosine monophosphate (cAMP) accumulation and phospholipids turnover and eventually PRL release, we have performed studies with MMQ cells. For this purpose, the levels of various G proteins (alpha o, alpha s, alpha i, alpha q and beta) and their mRNAs, measured by Western and Northern blots respectively, were correlated with intracellular cAMP accumulation in response to DA, VIP or DA plus VIP, and with inositol phosphates (IPx) formation in response to ANG II, DA or DA plus ANG II. This study shows that, when compared to normal pituitary tissue, the levels of alpha o, alpha o2 and alpha i3 were significantly decreased in MMQ cells; those of alpha o1, alpha i (alpha i1 + alpha i2), alpha s42 and alpha q were very low or undetectable while those of alpha s47 and beta were normal. DA was unable to inhibit basal PRL release and cAMP accumulation. VIP increased both cAMP accumulation and PRL release, while cAMP accumulation elicited by VIP could be suppressed by DA. BAY K 8644-induced PRL release also could be suppressed by DA.(ABSTRACT TRUNCATED AT 250 WORDS)

Efficacy of dobutamine in the failing transplanted heart.

Acute rejection often leads to severe myocardial failure and death. Surprisingly, no systematic study on the efficacy of beta-adrenergic pharmacologic agents have been reported to the present. Because of all the pathophysiologic alterations documented during rejection, we expected an inappropriate response to inotropic drugs, so we have questioned the value of dobutamine during those circumstances. Twelve dogs underwent orthotopic transplantation and were prepared with implantable devices for serial hemodynamic studies to be performed on the resting unanesthetized subject. Of this number, six dogs were studied while they were in immediate postoperative heart failure (3 hours after operation), and the same study was performed when myocardial failure secondary to rejection occurred (5 to 7 days). After basal state measurement, 5 and 10 micrograms.kg-1.min-1 of dobutamine were infused continuously, and the hemodynamic response during the two phases was compared. The baseline cardiac index in the immediate postoperative period was 1.4 +/- 0.4 L.min-1.m2 and 1.8 +/- 1.0 L.min-1.m2 during rejection, showing a similar degree of heart failure. Dobutamine (5 micrograms.kg-1.min-1) increased cardiac index by 97% 3 hours after transplantation and by 35% during rejection (p < 0.05). With 10 micrograms.kg-1.min-1 of dobutamine, the difference between increments was not significant (99% versus 79%). Raising the infusion rate of the drug to 15 and 20 micrograms.kg-1.min-1 during rejection increased cardiac index by 97% and 118%, respectively. Interestingly, no detrimental tachycardia occurred with this increased dosage. Heart failure secondary to acute rejection can therefore be improved by dobutamine.(ABSTRACT TRUNCATED AT 250 WORDS)

Efficacy of isoproterenol on the failing transplanted heart during early acute rejection.

Acute rejection often leads to severe myocardial failure and death. The beneficial hemodynamic effects of isoproterenol in improving immediate postoperative heart failure have prompted its routine use after transplantation. However, because of the physiopathological alterations documented during rejection, an inappropriate response of the graft to isoproterenol administration could be expected. Six dogs received orthotopic transplants and were prepared with implantable devices for serial hemodynamic studies. The studies were performed on the resting unanesthetized subject 3 hours after operation when transient heart failure was present and repeated when myocardial failure secondary to rejection occurred. After basal state measurement, various doses of isoproterenol were infused and the hemodynamic responses during each period were compared. During rejection, the hemodynamic response to 0.05 and 0.10 micrograms.kg-1.min-1 was significantly lower when compared with the response in the postoperative period. To achieve similar postoperative chronotropic and inotropic effects, 0.35 microgram.kg-1.min-1 of isoproterenol was necessary. Isoproterenol is therefore effective in controlling myocardial failure during acute rejection despite a reduced sensitivity of the sinoatrial node and myocardial tissue.

G protein modulation by estrogens.

Levels of various G protein subunits were assayed by immunoblot and densitometry, using specific antibodies, in anterior pituitaries and striata of female rats exposed to physiological or pharmacological modifications of ovarian hormone levels and, for comparison, in the same tissues of coeval male rats. Treatment of ovariectomized rats with 17 beta-estradiol 10 micrograms/rat/day for 5, 10 or 20 days induced a time-dependent rise in plasma prolactin (PRL) levels. While no change in G protein levels was observed in the striatum, estrogen treatment induced a significant reduction of all pituitary G protein levels except those of alpha i1, which remained unchanged, and of alpha s42, which increased in a time-dependent manner. A highly significant correlation was observed between pituitary alpha s42 values and plasma PRL levels. During the estrous cycle, pituitary values of alpha o, alpha i3 and alpha s47 were generally lower than those of ovariectomized rats, suggesting the existence of tonic inhibitory influence of circulating ovarian hormones. Pituitary levels of alpha o, alpha i1 and alpha s42 also showed a significant modulation during the various phases of the estrous cycle, and those of alpha o, alpha i3, alpha s47 and beta were significantly lower in female than in male rats. No significant effects of estrous cycle hormone variations or sex differences were observed in the values of striatum G proteins. In conclusion, these data clearly indicate that ovarian hormones, and particularly estrogens, have a significant and specific effect on pituitary G protein levels which may modulate the secretion of pituitary hormones such as PRL.

G proteins in normal rat pituitaries and in prolactin-secreting rat pituitary tumors.

It is still undetermined which GTP-binding (G) protein is involved in the regulation of prolactin (PRL) release and through which effector. This study shows that, when compared to normal pituitary tissue, the levels of alpha o protein were very low in dopamine (DA)-resistant, PRL-secreting pituitary tumors 7315a and MtTW15, while alpha o mRNA was present in the two tumors. In the MtTW15 tumor alpha i1, alpha i2 and alpha i3 levels were decreased while those of alpha s42 and alpha s47 were increased, and in the 7315a tumor alpha i2, alpha i3 and beta levels were decreased and those of alpha s47 increased. In an estrone-induced, DA-sensitive prolactinoma the levels of alpha i3 were greatly reduced. DA was unable to inhibit basal PRL release by 7315a and MtTW15 and basal cAMP accumulation by adenomatous and MtTW15 cells. Vasoactive intestinal peptide (VIP) increased both cAMP accumulation and PRL release by all cell preparations which could be suppressed by DA with adenomatous and 7315a but not with MtTW15 cells. These and previously published results provide circumstantial evidence that alpha o, alpha i1 and alpha i3 are all involved in the transduction of the DA inhibitory message while alpha s47 transduces cAMP activating messages and alpha s42 is responsible for the constitutive activation of L-type Ca2+ channels, adenylate cyclase and baseline PRL release.

Structural differences between dopamine D2 receptors present in a rat pituitary adenoma and in transplantable rat pituitary tumors 7315a and MtTW15.

We have investigated the structure of dopamine (DA) D2 receptors present in an estrone-induced, prolactin (PRL)-secreting, DA-sensitive adenoma and in two PRL-secreting and DA-insensitive transplantable tumors 7315a and MtTW15, in order to identify better the anomalies present in DA-resistant lactotrophs. D2 receptors were found in both a high- and a low-affinity state in adenomatous lactotrophs as shown by displacement studies with the agonist N-propylnorapomorphine (NPA), but only in the low-affinity state in the two DA-resistant tumors. Treatment with the alkylating agent N-ethylmaleimide induced a disappearance of the high-affinity state of the D2 receptor in the adenoma and a reduction in receptor concentration, but did not have any effect on the affinity of receptors present in DA-resistant tumors. Moreover, target size analysis and radiation inactivation studies of D2 receptors, using membranes preincubated with NPA and [3H]spiperone as ligand or using [3H]NPA as ligand on membranes preparations, have shown the presence of distinct structural differences between adenomatous and tumoral D2 receptors and between the two tumoral receptors themselves; these results suggest that the normal functional unit of the D2 receptor is a dimer associated with a guanine nucleotide-binding protein (G protein) subunit and that tumoral D2 receptors may exist in various polymeric forms unassociated with G proteins. The anomalies found to be present in tumoral D2 receptor complexes may be responsible for the insensitivity of these tumors to dopaminergic agonists' inhibitory activity on PRL release and tumor growth.

Regulation of Basal and stimulated prolactin release in prolactin-secreting rat pituitary tumors*.

Abstract To better understand the mechanisms of the inhibitory effects of dopamine on pituitary prolactin release, we have utilized an estrone-induced, benign and dopamine-sensitive rat pituitary adenoma and two malignant, transplantable and dopamine-resistant rat pituitary tumors, 7315a and MITW15. Enzymatically dispersed and Percoll purified cells obtained from the three tissues were incubated for 30 min in media with or without Na(+) and in the presence or the absence of dopamine and/or various prolactin releasers for evaluating the secretion of prolactin under baseline and experimental conditions. In some experiments, the cells were pretreated for 16 h with pertussis toxin to evaluate the eventual presence and role of pertussis toxin-sensitive G proteins. Dopamine inhibited baseline prolactin release by adenomatous lactotrophs in a Na(+)-dependent manner, but was totally inactive with 7315a and MtTW15 cells. The Ca(2+) channel agonist BAY K 8644 stimulated prolactin release with all three preparations and its effects were enhanced by a Na(+)-free medium. Dopamine antagonized the stimulatory effects of BAY K 8644 with adenomatous and 7315a cells only, even in the absence of Na(+). Pertussis toxin pretreatment significantly increased baseline prolactin release by adenomatous and MtTW15 cells and abolished dopamine inhibition of adenomatous lactotrophs baseline hormone release. BAY K 8644, TRH and vasoactive intestinal peptide, stimulated prolactin release by adenomatous cells and this effect was antagonized by dopamine in a pertussis toxin-sensitive manner. All prolactin releasers, except TRH, were effective also with 7315a cells, and its actions were not blocked by pertussin toxin. The stimulatory effects of BAY K 8644 and vasoactive intestinal peptide on 7315a cells were enhanced by pertussis toxin pretreatment. The results obtained with an almost pure preparation of adenomatous lactotrophs confirm the existence of a dual mechanism of dopamine inhibitory action on prolactin release: 1) a Na(+)-dependent action exerted on baseline, and 2) a Na(+)-independent action exerted on stimulated prolactin release. They also indicate that both actions are exerted through pertussis toxin-sensitive G proteins. Furthermore, our results show the presence in transplantable pituitary tumors 7315a and MtTW15 of multiple and diverse anomalies in the regulation of prolactin release probably due, at least partly, to anomalies of one or more G proteins and/or neurotransmitter receptors.

Selective deficiency of guanine nucleotide-binding protein Go in two dopamine-resistant pituitary tumors.

G proteins were quantitated by immunoblot in normal rat anterior pituitary, an estrone-induced pituitary adenoma, and in two transplantable pituitary tumors resistant to dopamine, 7315a and MtTW15. Antisera specific for the alpha o and beta subunits or the alpha i subunit of G proteins were tested with all preparations. While the alpha i and beta subunits were found to be present in variable concentrations in all preparations, the alpha o subunit was very low or undetectable in the transplantable tumors. These findings suggest that tumor resistance to the dopamine inhibitory actions on prolactin release and on tumor growth may be due to the deficiency of a Go protein.

Human placental LHRH receptor: agonist and antagonist labeling produces differences in the size of the non-denatured, solubilized receptor.

Membranes of human term placenta were labeled with several photosensitive and non-photosensitive analogues of LHRH. In both groups agonistic and antagonistic peptide structures were tested. The photolabeling amino acid azidophenylalanine was placed either in positions 2, 6 or 7. All compounds were specifically displacing iodinated Buserelin from human placental membranes and from rat pituitary membranes. The iodinated photolabels were displaced by cold Buserelin, some compounds however had a high non-specific binding. Photolabeling experiments on human placental membranes with radioactive photolabels produced all a band at 58,000 daltons in SDS-gel electrophoresis. Solubilization with a non-denaturing detergent and gel filtration produced a major radioactivity peak which was attributed to the LHRH receptor. All labeling experiments with agonist labels produced Kav's indifferent from each other but significantly different from the Kav's of antagonist labeled membranes. This result was confirmed with similar experiments carried out with radioactive Buserelin and a radioactive antagonist under non-photolytic conditions. It is therefore concluded that the placental LHRH receptor contains an LHRH binding component of 58,000 daltons but that the native receptor is composed of several proteins. It is also concluded that agonist occupation of the receptor induces the association of a further component which might be involved in the transmembrane signalling system. The agonist labeled, solubilized native LHRH receptor has a Stokes radius of 52 A and the same, antagonist labeled receptor a radius of 48 A.

Characterization of D2 dopamine receptors in dopamine-resistant prolactin-secreting rat pituitary tumors 7315a and MtTW15.

We have investigated the structure of D2 receptors present in two prolactin-secreting, dopamine-resistant, transplantable rat pituitary tumors, 7315a and MtTW15. These receptors specifically bind with high affinity the dopamine antagonist [3H]spiroperidol when membrane bound or solubilized by [3-(3-cholamidopropyl)-dimethyl-ammonio]-1-propane sulfonate 10 mM and are pharmacologically characterized as D2 type. Target-size analysis by radiation inactivation indicated a molecular mass of approximately 100,000 and 200,000 daltons for receptors present respectively in 7315a and MtTW15 tumors either membrane bound or solubilized. The minimal size of the D2 binding site was evaluated at 94,000 daltons by photoaffinity labeling with [125I]azido-N-(p-aminophenethyl)-spiperone followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A guanine nucleotide had no effect on the displacing potency of the agonist N-propylnorapomorphine evaluated with membrane-bound or solubilized receptors obtained from either tumor. These results suggest the absence or inactivation of a guanine nucleotide binding protein in the receptorial complex of these tumors. Thus, our data indicate that a structural anomaly is present in the D2 receptorial complex of these prolactin-secreting rat pituitary tumors, which may be responsible for their resistance to the inhibitory effects of dopamine.

Tracheal growth after resection and anastomosis in puppies.

Tracheal morphology, morphometric changes, and growth and histologic changes were studied in puppies submitted to tracheal resection and anastomosis. Fifteen mongrel puppies about 12 weeks old and weighing on an average 5.5 kg were operated under general anesthesia using fluothane. A median cervicotomy incision was made in ten puppies (experimental group, EG) and the proximal 14 tracheal rings were resected (average length 5.08 cm or about 35% to 38% of total tracheal length). One layer anastomosis was done using vicryl 4.0 maintaining the average tension of 1,450 g. Five puppies (control group, CG) were submitted to tracheal transection and anastomosis and the following parameters were studied. Tracheal morphology the trachea of the EG was a rounded triangle whereas in the CG it was oval in shape, there was increase in the intercartilageneous spaces in the EG, no granulation tissue was present, two mucous webs were seen in the EG and one in the CG. Morphometric changes average tracheal length EG 13 cm, CG 17.7 cm, intercartilagenous space EG 3.08 mm, CG 1.3 mm, intercricothyroid space EG 1.2 cm, CG 0.53 cm, sagittal and transverse tracheal thickness at the anastomosis EG 2.6 and 3.3 mm, CG 2 and 1.5 mm, sagittal and transverse diameter reduced on an average 2 mm in EG. Histology Moderate fibrosis was found at the level of anastomosis with no modification of chondrocytes at the cartilagenous rings in the EG. Even with high anastomotic tension, the dogs had normal tracheal growth without stenosis; the sagittal and transverse growth at the anastomosis in the EG was 90% and 85%, respectively, when compared with the CG.

Photolysis and ozonolysis of desmosine and elastolytic peptides.

This paper describes the combined effects of photolysis and ozonolysis upon the main interchain crosslinks of elastin, desmosine and isodesmosine. Photolysis of purified (iso)desmosines in solution leads to the cleavage of the pyridinium rings to give lysine and an analogue of glutaconic aldehyde which is deformylated to a stable compound absorbing at 242 nm. This photoproduct is subsequently fragmented by ozone into glutamic, alpha-aminoadipic and possibly alpha-amino-delta-oxocaproic acids. However the yields of these different compounds are very low because we observed that numerous competing side reactions (polymerisation, recyclisation) accompany the photoozonolytic decomposition of the pyridinium rings of free (iso)desmosines. The results are clearer when reticulated elastolytic peptides are photoozonolysed. The (iso)desmosines, covalently linked in these peptides, are cleaved into lysine, glutamic and alpha-aminoadipic acids (in a ratio 2:1) with yields corresponding to 80-90% of those expected from the decomposition of the (iso)desmosines originally present in the peptide fraction. We have also observed that ozonolysis alone degraded another crosslink present in these peptides, the aldol condensation product, resulting in the production again of glutamic and alpha-aminoadipic acids in amounts consistent with the known concentrations of this particular crosslink in elastin. Finally we noted that the complete photoozonolytic degradation of the (iso)desmosines present in a semi purified reticulated elastolytic fraction resulted in a shift of the size distribution of these peptides toward lower values. It is not certain however that this shift, indicative of a freeing of the polypeptide chains from their original three dimensional network, is due uniquely to the cleavage of the (iso)desmosines. Indeed we have observed that tyrosine and phenylalanine were also degraded during photoozonolysis. Not knowing the mechanism of this degradation it is impossible to rule out the possibility that concomitant cleavages of peptide bonds did occur.