Biomedical subjects

T Takao

Publications and source records attributed to T Takao.

At least 199 records · Page 11Linked to original sources

PubMed1990

Identification of interleukin-1 receptors in mouse testis.

The cytokine interleukin-1 (IL-1) has been reported to alter reproductive functions through both effects in brain and direct actions at the level of the gonads. To further define the role of IL-1 at the gonads, we have used 125I-labeled recombinant human IL-1 to identify and characterize IL-1 receptors in crude membrane preparations of mouse (C57BL/6) testis and to study the distribution of IL-1-binding sites using autoradiography. In preliminary homogenate binding and autoradiographic studies, [125I]IL-1 alpha showed significantly higher specific binding than [125I]IL-1 beta. Thus, [125I]IL-1 alpha was used in all subsequent assays. The binding of [125I]IL-1 alpha was linear over a broad range of membrane protein concentrations, saturable, and reversible, and on Scatchard analysis revealed an equilibrium dissociation constant (Kd) of 82 +/- 4 pM and a maximum number of binding sites (Bmax) of 10.8 +/- 1.5 fmol/mg protein. In competition studies, IL-1 alpha, IL-1 beta, and a weak IL-1 beta analog IL-1 beta+ inhibited [125I]IL-1 alpha binding to mouse testis in parallel with their relative bioactivities in immune assays, with inhibitory binding affinity constant (Ki) values of 14.2 +/- 1.7, 88.8 +/- 5.7, and 7183.3 +/- 603 pM, respectively; rat/human CRF and human tumor necrosis factor showed no effect on [125I]IL-1 alpha binding. In autoradiographic studies, IL-1 receptors were heterogeneously distributed, with highest densities present in the luminal border of the epididymis and interstitial areas of the testis. After hypophysectomy, the testes were significantly atrophied, and homogenate binding and autoradiographic studies showed that while the total number of binding sites per testis was significantly decreased in hypophysectomized mice in proportion to the reduction in testicular mass, there was no apparent change in the relative density of IL-1 receptors. These data provide the first identification of IL-1 receptors in testis and provide further support for a physiological role for IL-1 to alter reproductive functions through a direct effect at the gonads.

Animals↗

PubMed1990

Interleukin-1 receptors in mouse brain: characterization and neuronal localization.

The cytokine interleukin-1 (IL-1) has a variety of effects in brain, including induction of fever, alteration of slow wave sleep, and alteration of neuroendocrine activity. To examine the potential sites of action of IL-1 in brain, we used iodine-125-labeled recombinant human interleukin-1 [( 125I]IL-1) to identify and characterize IL-1 receptors in crude membrane preparations of mouse (C57BL/6) hippocampus and to study the distribution of IL-1-binding sites in brain using autoradiography. In preliminary homogenate binding and autoradiographic studies, [125I]IL-1 alpha showed significantly higher specific binding than [125I]IL-1 beta. Thus, [125I]IL-1 alpha was used in all subsequent assays. The binding of [125I]IL-1 alpha was linear over a broad range of membrane protein concentrations, saturable, reversible, and of high affinity, with an equilibrium dissociation constant value of 114 +/- 35 pM and a maximum number of binding sites of 2.5 +/- 0.4 fmol/mg protein. In competition studies, recombinant human IL-1 alpha, recombinant human IL-1 beta, and a weak IL-1 beta analog. IL-1 beta +, inhibited [125I]IL-1 alpha binding to mouse hippocampus in parallel with their relative bioactivities in the T-cell comitogenesis assay, with inhibitory binding affinity constants of 55 +/- 18, 76 +/- 20, and 2940 +/- 742 pM, respectively; rat/human CRF and human tumor necrosis factor showed no effect on [125I]IL-1 alpha binding. Autoradiographic localization studies revealed very low densities of [125I]IL-1 alpha-binding sites throughout the brain, with highest densities present in the molecular and granular layers of the dentate gyrus of the hippocampus and in the choroid plexus. Quinolinic acid lesion studies demonstrated that the [125I]IL-1 alpha-binding sites in the hippocampus were localized to intrinsic neurons. After hypophysectomy, homogenate binding and autoradiographic studies showed that there was no apparent change in the relative density of IL-1 receptors in the hippocampus. The identification of IL-1 receptors in brain with characteristics similar to IL-1 receptors in immune and neuroendocrine tissues provides further support for a physiological role for IL-1 to regulate central nervous system activity.

Animals↗

PubMed1990

Effect of hypertonic saline on the corticotropin-releasing hormone and arginine vasopressin content of the rat pituitary neurointermediate lobe.

The changes in the levels of corticotropin-releasing hormone (CRH) and arginine vasopressin (AVP) in the neurointermediate lobe of the pituitary (NIL) following hypertonic saline administration were examined in rats. The plasma osmotic pressure in rats receiving 2% NaCl for 8 days was greatly increased. Plasma AVP concentration in rats receiving 2% NaCl for 8 days were significantly higher than in control rats (566% of the control level). Plasma corticosterone was significantly higher in the saline-treated rats than in controls, whereas plasma ACTH was not significantly different. The pituitary ACTH concentration was much higher in the saline-treated rats than in controls. CRH in the NIL was increased significantly by saline treatment (419% of the control concentration), whereas the CRH in the paraventricular nucleus and median eminence of control and saline-treated rats did not differ significantly. The AVP in the NIL fell greatly in saline treated rats. The extract from both control and saline-treated rats showed a major peak for immunoreactive CRH, with a retention time identical to that of rat CRH. However, the peak was much higher in the extract from saline-treated rats. The immunoreactive AVP peak was greatly reduced in saline-treated rats. These results suggest that hypertonic saline administration increases the CRH in the NIL and causes AVP hypersecretion and/or hyperfunction of magnocellular-NIL CRH might be responsible for pituitary-adrenal stimulation in saline-treated rats.

Adrenal Cortex Hormones↗

PubMed1990

Effects of hyperosmotic stimulation and adrenalectomy on vasopressin mRNA levels in the paraventricular and supraoptic nuclei of the hypothalamus: in situ hybridization histochemical analysis using a synthetic oligonucleotide probe.

The effects of salt loading and adrenalectomy on arginine vasopressin (AVP) mRNA levels in the paraventricular nucleus (PVN) and the supraoptic nucleus (SON) of the hypothalamus were studied by semiquantitative in situ hybridization histochemistry, using a synthetic oligonucleotide probe and a computer-assisted image analysis system. Salt loading (2% NaCl) for 7 days produced marked increases in AVP mRNA levels in the magnocellular neurons of the PVN, SON, and accessory nuclei. Adrenalectomy caused an increase in AVP mRNA expression in the magnocellular part of the PVN and the expansion of hybridization signals into its medial parvocellular region, where the cell bodies of corticotropin-releasing hormone (CRH) neurons are located. No apparent alteration of AVP mRNA levels was observed in the SON following adrenalectomy. These results indicate that hyperosmotic stimulation and the loss of circulating glucocorticoids had differential effects on AVP gene expression in the PVN and SON, and that the magnocellular PVN and SON neurons responded in different manners to the loss of feedback signals.

Adrenalectomy↗

PubMed1990

Combined anterior pituitary function test using CRH, GRH, LH-RH, TRH and vasopressin in patients with non-functioning pituitary tumors.

We examined 8 normal subjects and 16 patients with non-functioning pituitary tumors with a combined anterior pituitary test to evaluate the clinical usefulness of the test. Diagnoses included 9 of chromophobe adenoma, 3 of craniopharyngioma, 2 of Rathke's cleft cyst, and 1 each of intrasellar cyst and tuberculum sella meningioma. All subjects received hypothalamic releasing hormones: 1 micrograms/kg corticotropin releasing hormone (CRH), 1 micrograms/kg growth hormone releasing hormone (GRH), 500 micrograms thyrotropin-releasing hormone (TRH), 100 micrograms luteinizing hormone releasing hormone (LH-RH), and a relatively small dose (5 mU/kg) of lysine vasopressin (LVP). In the normal subjects, the addition of LVP potentiated the secretion of adenocorticotropic hormone (ACTH) induced by CRH, but had no significant effect on the secretion of other anterior pituitary hormones. In the combined test with 5 releasing hormones, the plasma ACTH and cortisol responses were not impaired in the majority of the patients before pituitary surgery. Serum thyroid-stimulating hormone (TSH), prolactin (PRL) and follicle-stimulating hormone (FSH) responses were not impaired in 82%, 70% and 67% of the patients, respectively, while the serum LH and GH responses were impaired in 67% and 73% of the patients, respectively. Following pituitary surgery, responses of these hormones to combined testing were similarly impaired in more than 75% of the patients. These results indicate that plasma ACTH, cortisol and serum TSH responses are fairly good before pituitary surgery but are impaired significantly after surgery. No subjects experienced any serious adverse effects related to the testing. These results suggest that combined testing with hypothalamic hormones is a convenient and useful method for evaluating pituitary function.

Adult↗

PubMed1990

[A case report of synchronous double cancer of the lung and esophagus].

A 69-year-old man was admitted to our department with complaint of difficult swallowing. Upper gastrointestinal examination showed esophageal cancer and squamous cell carcinoma of the middle thoracic esophagus was revealed histologically. Chest X-ray showed the atelectasis of the B-3 region of right upper lobe of the lung. Further examinations revealed histologically squamous cell carcinoma growing from B-3 bronchus. Finally, he was diagnosed as synchronous double carcinoma of the lung and the esophagus. Simultaneous operation was performed on May 20, 1988. First, right upper lobectomy of the lung and wedged resection of right main bronchus were performed. Esophagectomy and antesternal esophageal reconstruction using the gastric tube was added. The postoperative course was satisfactory without any severe complications. The synchronous double cancer of the lung and the esophagus is rare. Our case indicates that simultaneous resection of both cancer and esophageal reconstruction can be safely performed.

Aged↗

PubMed1990

[Double chambered right ventricle: surgical experience in a 60-year-old woman].

The patients reported as double chambered right ventricle are mainly children. A 60-year-old woman had been pointed out for her systolic cardiac murmur without any symptom. She was admitted to our hospital for her gradual onset of fatigue, lassitude. Cardiac catheterization data revealed a 105 mmHg peak-to-peak gradient within the right ventricular cavity with normal pulmonary pressures [20/5 (12) mmHg]. Right ventricular end-diastolic pressure was 4 mmHg. Right ventriculogram demonstrated double chambered right ventricle. Electrocardiogram showed neither right ventricular hypertrophy nor upright T wave in V3R. In order to release intraventricular pressure gradient and her symptom, the anomalous muscle bundles were resected through both the right atrium and the pulmonary artery. Surgical repair without right ventriculotomy is suitable for such an elder patient with double chambered right ventricle whose ventricular function may decrease.

Age Factors↗

PubMed1989

Identification of a disaccharide (Xyl-Glc) and a trisaccharide (Xyl2-Glc) O-glycosidically linked to a serine residue in the first epidermal growth factor-like domain of human factors VII and IX and protein Z and bovine protein Z.

We have recently described a unique trisaccharide linked to a serine residue in the first epidermal growth factor-like domains of bovine blood coagulation factors VII (Ser-52) and IX (Ser-53) (Hase, S., Kawabata, S., Nishimura, H., Takeya, H., Sueyoshi, T., Miyata, T., Iwanaga, S., Takao, T., Shimonishi, Y., and Ikenaka, T. (1988) J. Biochem. (Tokyo) 104, 867-868). The sugar chain identified in these clotting factors consists of 1 mol of hexose (glucose (Glc] and 2 mol of pentose (xylose (Xyl]. We report here that human factors VII and IX and protein Z and bovine protein Z also contain such carbohydrate moieties linked to a serine residue at the same position found in bovine factors VII and IX. A glycopeptide derived from each of these proteins was subjected to amino acid sequence and component sugar analyses and fast atom bombardment mass spectrometric analysis. The results indicate that the glycopeptide derived from human factor IX contains 1 mol each of Glc and Xyl. The reducing end of this disaccharide was identified as Glc by analyzing the disaccharide generated by hydrazinolysis. In contrast, human factor VII and protein Z yielded two different glycopeptides which contained Glc and Xyl at molar ratios of 1:1 and 1:2, respectively, suggesting microheterogeneity of these O-linked sugar chains. Bovine protein Z glycopeptide contained 1 mol of Glc and 2 mol of Xyl. These sugar compositions were confirmed by analyses of the intact proteins. In relation to the trisaccharide sugar chain previously discovered in bovine factors VII and IX, these findings indicate the existence of a Xyl2-Glc-Ser and a Xyl-Glc-Ser structure in the first epidermal growth factor-like domains of human factors VII and IX and protein Z in addition to that of bovine protein Z. Whether these carbohydrate moieties contribute to the biological activities of these proteins is unknown.

Amino Acid Sequence↗

PubMed1989

Sugar sequences of allergenically active oligosaccharide alcohols isolated from a large-molecular-size sea squirt antigen termed H-antigen.

O-Glycosidically linked oligosaccharide chains were liberated as oligosaccharide alcohols (HPG-beta 2) by beta-elimination treatment from a glycopeptide fraction (HPG, Mr ca. 10(5)) that was isolated from a Pronase digest of a large-size sea squirt antigen termed H-antigen (Mr 10(6)-2 X 10(7)). After HPG-beta 2 was divided into a neutral (HPG-beta 2-N) and an acidic (HPG-beta 2-A) fraction, 11 neutral saccharitols were isolated from HPG-beta 2-N, and their sugar sequences were determined by methylation/GC-MS, FAB-MS, and/or DI/EI-MS. By skin tests specific to patients with sea squirt allergy, it was found that six saccharitols were allergenically active but five were inactive. Furthermore, active saccharitols were all of branched structure containing two nonreducing terminal GalNAc (t-GalNAc) residues, whereas the inactive saccharitols contained one or no t-GalNAc. Since the allergic reaction in skin should depend on the crosslinking of IgE antibodies by certain allergens carrying two or more epitopes, three types of N-acetylgalactosaminyl disaccharide units, GalNAc1----2Fuc1----, GalNAc1----3/4GalNAc1----, and GalNAc1----3/4GlcNAc1----, were nominated as the major components of the allergy-specific epitopes in the active saccharitols. Such multiplicity of the epitope structures might reflect the polyclonality of the specific IgE antibodies.

Allergens↗

PubMed1989

Induction of nonspecific killer cells by delayed-type hypersensitivity against soluble protein antigens in murine peritoneal cavities.

We induced nonspecific killer cells in the local site of delayed-type hypersensitivity against keyhole limpet hemocyanin or ovalbumin. Delayed-type hypersensitivity was induced in the peritoneal cavities of mice, and peritoneal exudate cells (PEC) were collected. These PEC were found to have killer activity toward SP2 and YAC-1 cells (target cells susceptible to natural killer cells) by 4-h 51Cr-release assays. The induction of killer activity in PEC was observed in parallel with the eliciting of delayed-type hypersensitivity in the peritoneal cavity, in which the killer activity was maximum 24-48 h after the antigen challenge, but was not induced in nu/nu mice and was induced in an antigen-specific way. These killer cells did not adhere to nylon wool and had Thy1 and asialo-GM1 antigens on their surfaces. Their precursor cells were also asialo-GM1-positive. These findings indicate that the killer cells probably belong to the NK cell lineage. Results of tumor challenge experiments showed that these killer cells had an antitumor effect in vivo as well as in vitro.

Animals↗

PubMed1989

Antimicrobial peptides, isolated from horseshoe crab hemocytes, tachyplesin II, and polyphemusins I and II: chemical structures and biological activity.

Tachyplesin is an antimicrobial peptide recently found in the acid extract of hemocytes from the Japanese horseshoe crab (Tachypleus tridentatus) [Nakamura, T. et al. (1988) J. Biol. Chem. 263, 16709-16713]. In our continuing studies on the peptide, we have found an isopeptide, tachyplesin II, and also polyphemusins I and II in hemocytes of the American horseshoe crab (Limulus polyphemus). The complete primary structures of these peptides, which are very similar to that of the previously isolated peptide, now named tachyplesin I, were determined to be as follows: (Table: see text). The isopeptide, tachyplesin II, consists of 17 residues with a COOH-terminal arginine alpha-amide. On the other hand, both polyphemusins I and II were found to contain 18 residues due to an additional Arg residue at the NH2-terminal end as well as a COOH-terminal arginine alpha-amide. The disulfide linkages for polyphemusin I consisted of two bridges between Cys-4 and Cys-17 and between Cys-8 and Cys-13, which was identical to in the case of tachyplesin I. Moreover, all of these peptides inhibited the growth of not only Gram-negative and -positive bacteria but also fungi, such as Candida albicans M9. Furthermore, complex formation between these peptides and bacterial lipopolysaccharides was also observed in a double diffusion test. These results suggest that tachyplesins and polyphemusins are probably located in the hemocyte membrane, where they act on antimicrobial peptides as a self-defense mechanism in the horseshoe crab against invading microorganisms.

Alkylation↗

PubMed1989

Abnormalities in the hypothalamo-pituitary-adrenal axis in spontaneously hypertensive rats during development of hypertension.

Abnormalities in the hypothalamo-pituitary-adrenal axis in spontaneously hypertensive rats (SHR) during development of hypertension were investigated using in vivo and in vitro methods. Plasma ACTH responses to hemorrhage and ether stress were significantly smaller in 7-week-old SHR than in age-matched Wistar-Kyoto rats (WKY), while plasma corticosterone baseline levels and its response to stress were greater in SHR than in WKY. There was no significant difference in the plasma ACTH response to ether stress between bilaterally adrenalectomized SHR and WKY replaced with a 25% corticosterone pellet for 6 days. Adrenalectomy prevented the development of hypertension in SHR; however, corticosterone replacement restored hypertension. Plasma ACTH showed a smaller response to iv CRH injection in SHR than in WKY, while the ACTH response to arginine vasopressin was not different between SHR and WKY. CRH concentrations in the median eminence, posterior pituitary, and cerebral cortex were lower in SHR than in WKY, while the CRH concentration in the median eminence was not different in SHR and WKY when they were adrenalectomized with or without corticosterone replacement. Basal in vitro CRH release from hypothalamic tissue was reduced in SHR, while CRH release in response to 56 mM KCl was not different in SHR and WKY. These results suggest that adrenocortical function is enhanced in young SHR, that reduced ACTH response to stress and exogenous CRH in SHR may be ascribed to higher plasma corticosterone levels, and that corticosterone is essential for the development of hypertension in SHR.

Adrenalectomy↗

PubMed1989

Cushing's syndrome due to huge nodular adrenocortical hyperplasia with fluctuation of urinary 17-OHCS excretion.

A 51-yr-old male patient with Cushing's syndrome due to huge nodular adrenocortical hyperplasia is described. Urinary 17-OHCS was not suppressed by a high dose of (8 mg) dexamethasone and showed rather a tendency to paradoxical response. There was no response to metyrapone. Plasma cortisol showed a hyperresponse to insulin-induced hypoglycemia and a rapid response to corticotropin releasing hormone-lysine vasopressin (CRH-LVP) administration without an obvious ACTH response. Plasma cortisol responded to synthetic ACTH. Urinary 17-OHCS did not show parallel changes with plasma cortisol. These results and computerized tomography data suggested huge multiple nodular adrenocortical hyperplasia, which was confirmed later by surgery. The left and right adrenal glands weighed 105 and 45 g, respectively. Hyper-reaction of the adrenal gland to a small change in plasma ACTH or "unknown factors" may cause not only the discrepancy between cortisol and ACTH response but also the development of autonomous nodules in the adrenal gland.

17-Hydroxycorticosteroids↗

PubMed1989

Effect of acute ether or restraint stress on plasma corticotropin-releasing hormone, vasopressin and oxytocin levels in the rat.

Ether and restraint stress-induced peripheral plasma corticotropin releasing hormone (CRH), arginine vasopressin (AVP), oxytocin (OXY) and adrenocorticotropin (ACTH) levels were measured by radioimmunoassays. Plasma CRH, AVP, OXY and ACTH rose to approximately twice the level of control rats 2 min after the onset of a 1-min exposure to ether. Plasma CRH rose further 5 min after the onset of ether stress, while plasma AVP and OXY returned to the baseline levels at 5 min. Plasma CRH, OXY and ACTH showed significant elevation 2 min after the onset of restraint stress, while plasma AVP did not show a significant change. Plasma OXY and ACTH rose further 5 min after the onset of restraint stress, whereas plasma CRH returned to baseline levels. CRH and OXY concentrations in the hypothalamic median eminence decreased 5 min after the onset of ether exposure and restraint, while the AVP concentration did not differ from control levels. The results, including the discrepancy between plasma CRH and ACTH 5 min after stress, suggest that CRH in the peripheral plasma is derived from both hypothalamic and extrahypothalamic tissues. The levels of stress-induced CRH in the peripheral plasma were sufficient to stimulate ACTH release. These results suggest that ether and restraint stress elevate plasma CRH shortly after the onset of the stress, and that this elevation in the plasma CRH level is at least partly responsible for stress-induced ACTH secretion.

Adrenocorticotropic Hormone↗

PubMed1989

[Idiopathic chylothorax in an adult--a case report and a review of 14 cases in Japan].

A 30-year-old female with an idiopathic chylothorax who underwent successful surgical treatment was reported. The patient was referred to our hospital because of massive right pleural effusion found by chance on the X-ray. Thoracocentesis revealed chyle but the etiology was not known by examinations. Conservative treatments such as fasting, intravenous hyper-alimentation and a continuous drainage were performed for 10 weeks without remarkable improvement. A right thoracotomy revealed nothing abnormal except for the oozing of lymph from mediastinal pleura, which was sutured by 3-0 nylon. Postoperative course was uneventful and no recurrent pleural effusion has been recognized for 3 years since then. Adult chylothorax with unknown etiology like this is rare and only 13 cases have been reported in Japan. Such "idiopathic chylothorax" cases, including our case, were reviewed and discussed.

Adult↗

PubMed1988

Tachyplesin, a class of antimicrobial peptide from the hemocytes of the horseshoe crab (Tachypleus tridentatus). Isolation and chemical structure.

A cationic peptide, designated tachyplesin, was isolated from acid extracts of horseshoe crab (Tachypleus tridentatus) hemocyte debris. It consists of 17 residues and the structure determined by Edman degradation is: (formula; see text) The carboxyl-terminal end of this peptide was identified as arginine alpha-amide, and the whole sequence including the alpha-amide was also confirmed by fast atom bombardment mass spectrometry, indicating a mass value of 2263. Tachyplesin inhibits growth of both Gram-negative and -positive bacteria at low concentrations and formed a complex with bacterial lipopolysaccharide. Tachyplesin seems likely to act as antimicrobial peptide for self-defense in the horseshoe crab against invading microorganisms.

Amino Acid Sequence↗

PubMed1988

Bovine factor VII. Its purification and complete amino acid sequence.

A modified method for purification of blood clotting factor VII from bovine plasma was developed, and its complete amino acid sequence was established. The isolated factor VII was activated with factor XIIa, and the resulting two-chain factor VII (factor VIIa) was reduced and S-pyridylethylated or S-aminoethylated. The amino acid sequences of the S-alkylated heavy and light chains were determined by sequencing the fragments obtained from enzymatic and chemical cleavages. Fast atom bombardment mass spectrometry was also used to establish the COOH-terminal sequence of the heavy chain. The light chain consists of 152 residues with one carbohydrate chain at Asn145, and 11 gamma-carboxyglutamic acid residues are found within the NH2-terminal 35 residues. The light chain contains 0.2-0.3 mol of beta-hydroxyaspartic acid/mol of protein, indicating that an aspartic acid residue in bovine factor VII is incompletely hydroxylated. Moreover, a pentapeptide, Ala-Ser*-Ser-Pro-Cys (positions 51-55), isolated from an enzymatic digest of the light chain, contained an unknown serine derivative, but its structure is still unclear. On the other hand, the heavy chain is composed of 255 residues and one asparagine-linked carbohydrate chain at Asn203. Bovine factor VII, with a total of 407 residues, has 71% sequence identity with the human molecule (406 residues) predicted from the cDNA sequence (Hagen, F. S., Gray, C. L., O'Hara, P., Grant, F. J., Saari, G. C., Woodbury, R. G., Hart, C. E., Insley, M., Kisiel, W., Kurachi, K., and Davie, E. W. (1986) Proc. Natl. Acad. Sci. U. S. A. 83, 2412-2416).

Amino Acid Sequence↗

PubMed1988

Substitution of gamma Arg-275 by Cys in an abnormal fibrinogen, "fibrinogen Osaka II". Evidence for a unique solitary cystine structure at the mutation site.

In an abnormal fibrinogen with impaired fibrin monomer polymerization designed as fibrinogen Osaka II, we have identified substitution of Arg by Cys at position 275 of the gamma chain. This Cys is linked to a free cysteine molecule by a disulfide link as evidenced by fast atom bombardment mass spectrometry. This finding was supported by identification of a single cysteine released from isolated abnormal fragment D1 upon reduction. This unique cystine structure at the mutation site has not been reported heretofore in any abnormal protein including fibrinogen. The substitution may well perturb the structure required for fibrin monomer polymerization, specifically that assigned to the carboxyl-terminal D domain of fibrinogen. Indeed, isolated fragment D1 with the Cys substitution failed to inhibit thrombin-mediated clotting of normal fibrinogen and normal fibrin monomer polymerization, while normal fragment D1 inhibited them markedly. Our data seem to provide supporting evidence that the putative polymerization site(s) assigned to the D domain of fibrinogen may be structure-dependent, including the carboxyl-terminal segment of the gamma chain as well as a contiguous region that contains the gamma 275 residue.

Adolescent↗