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The synthesis and structure of the title compound, (C(24)H(20)P)(2)[Ge(C(2)O(4))(3)], are reported. The PPh(4)(+) cations in the structure form infinite zigzag chains in which the P.P distances alternate between 6.229 (1) and 7.118 (1) A, and the P.P.P angle is 131.4 (1) A. The shorter P.P distance is associated with a sixfold phenyl embrace. However, the longer P.P distance is associated with both phenyl-phenyl interactions and interactions between the cations and a twofold symmetric [Ge(C(2)O(4))(3)](2-) anion. In the cation-anion interactions, the P.O distance is 4.444 (2) A, the O.P-C(distal) angle is 175.0 (1) degrees and the shortest H.O distances are 2.74 and 3.09 A.

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The compounds (Me4N)[A(M(SC(O)Ph)3)2] (A = K, M = Cd (2); A = Na, M = Hg (3); and A = K, M = Hg (4)) were synthesized by reacting the appropriate metal chloride with A+PhC(O)S- and Me4NCl in the ratios 1:3:1 and 2:6:1. The structures of these compounds were determined by single-crystal X-ray diffraction methods. All the compounds are isomorphous, isostructural, and crystallized in the space group P1 with Z = 1. Single-crystal data for 2: a = 106670(2) A, b = 111522(2) A, c = 119294(2) A, alpha = 71782(1) degrees, beta = 85208(1) degrees, gamma = 69418(1) degrees, V = 126140(4) A3, Dcalc = 1528 g cm-3. Single-crystal data for 3: a = 10840(2) A, b = 10946(4) A, c = 12006(3) A, alpha = 7218(2) degrees, beta = 8675(2) degrees, gamma = 6743(2) degrees, V = 12493(6) A3, Dcalc = 1756 g cm-3. Single-crystal data for 4: a = 104780(1) A, b = 112563(2) A, c = 119827(2) A, alpha = 71574(1) degrees, beta = 85084(1) degrees, gamma = 70705(1) degrees, V = 126523(3) A3, Dcalc = 1755 g cm-3. In the [A(M(SC(O)Ph)3)2]- anions, each M(II) atom is bonded to three thiobenzoate ligands through sulfur atoms, giving a trigonal planar MS3 geometry. The carbonyl oxygen atoms from the two [M(SC(O)Ph)3]- anions are bonded to the alkali metal atom, providing an octahedral environment. Solution metal NMR studies showed the concentration-dependent dissociation of the alkali metal ions in the trinuclear anions.

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BACKGROUND: Acute left-sided colonic obstruction is a surgical emergency whose management is controversial. Because experience using expandable metal stents for relief of this type of obstruction is limited, we evaluated their effectiveness, feasibility, safety, and outcome. METHODS: Twenty-five patients with acute colorectal obstruction underwent placement of various metal stents under fluoroscopic and endoscopic guidance. On an intent-to-treat basis, stents were placed for decompression before one-stage surgical resection in 10 patients and palliatively in 15 patients. Two preoperative patients had unresectable malignant disease, and stents were left for palliation resulting in 17 palliative and 10 preoperative patients for analysis. RESULTS: Stent placement was technically successful in 94% of patients. Overall effectiveness in relieving obstruction was 85% (palliative 82%, preoperative 90%). In the palliative group, stent duration ranged from 2 to 64 weeks (mean 17.3 weeks). Major complications occurred in 7 patients (30%). CONCLUSIONS: Expandable metal stents are a feasible, effective adjunct and alternative to surgery for acute colorectal obstruction.

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The idiopathic inflammatory bowel diseases, ulcerative colitis and Crohn's disease, are chronic disorders that appear to arise from an aberrant interaction of environmental, genetic, and immunologic factors. The aim of this study was to examine the immune reactivity of a spontaneously colitic mouse strain, C3H/HeJBir, to epithelial, food, and enteric bacterial Ags. Serum Ab responses of colitic C3H/HeJBir and noncolitic parental C3H/HeJ mice were measured by enhanced chemiluminescence Western blotting. No reactivity to epithelial or food Ags was detected. However, the sera from C3H/HeJBir mice had a reproducible banding pattern on Western blot to bacterial Ags, whereas sera from C3H/HeJ mice did not. Only a small, highly selected number of enteric bacterial Ags were recognized. There were major differences in the degree of recognition of different bacterial strains, marked by remarkably few Abs to Ags of the major anaerobes of the bacterial flora. The serum Abs detected on immunoblot were primarily IgG2a, suggesting a Th1 response. Comparison of sera reactivity to histopathologic severity showed an inverse relationship: one third of young C3H/HeJBir mice during the peak of colitis produced Abs to bacterial Ags, while later in life, when the colitis had resolved, 96% produced Abs. These data are consistent with an abnormal immune reactivity to enteric bacterial flora in C3H/HeJBir mice, a reactivity that is highly selective considering the abundant bacterial Ags present in the colon lumen. We postulate that this reactivity plays a role in the pathogenesis of colitis in these mice.

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OBJECTIVE: The purpose of this prospective study was to evaluate the efficacy of expandable metal stents for colonic decompression in patients presenting with acute malignant obstruction and to describe the associated radiographic findings. SUBJECTS AND METHODS: Using both fluoroscopic and endoscopic guidance, we placed expandable metal stents within the colons of 13 patients presenting with acute malignant obstruction. Stents were placed in four patients to permit a standard bowel cleansing before surgical resection with primary anastomosis. In the other nine patients, stents were placed for palliation of nonresectable tumors, obviating colostomy. Outcomes and complications were analyzed. The radiologic aspects of procedural planning, stent placement, assessment after placement, and detection of complications were evaluated. RESULTS: Of the four surgical candidates who were successfully resected with primary anastomosis, two received incomplete bowel cleansing because of stent migration with recurrent obstruction. Eight of the nine patients who had stents placed for palliation of nonresectable tumors had relief of acute obstruction. Complications in this group included two perforations, one that required immediate colostomy and one that was self-limited and conservatively treated. Two other patients of the eight developed early stent obstruction, requiring colostomy in one. The other patient who had a stent placed for palliation of a nonresectable tumor declined further treatment. Contrast-enhanced enema examination proved useful in assessing the suitability of lesions for stent decompression, directing the choice of stent type and the most appropriate position for the patient during the stent placement. Immediately after stent placement, plain abdominal radiographs and water-soluble contrast enema examinations helped us verify stent position and patency. CONCLUSION: These results suggest that placement of expandable metal stents in patients presenting with acute, malignant colonic obstruction is a viable alternative to colostomy and can potentially decrease morbidity and mortality. Radiologic assessment before, during, and after stent placement plays an integral role in the management of patients undergoing stent decompression of the colon.

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PURPOSE: The aim of this study was to identify the overall long-term causes of death in a large series of patients who were undergoing proctocolectomy with ileal pouch-anal anastomosis (IPAA). METHODS: Records of patients who underwent proctocolectomy with IPAA at the Mayo Clinic affiliated hospitals between January 1981 and October 1994 were reviewed to determine overall mortality, cause, and timing of death. RESULTS: A total of 1,603 patients underwent proctocolectomy with IPAA reconstruction (1,407 for chronic ulcerative colitis (CUC), 187 for familial polyposis (FAP), and 9 for other diagnoses). Thirty-two patients have died, with an overall mortality rate of 2 percent. Mean age at time of death was 40 (23-60) years. There was no significant difference in overall mortality between patients with CUC and patients with FAP. Three deaths occurred postoperatively (0.2 percent) because of pulmonary embolism, perforated gastric ulcer, and subarachnoid hemorrhage. Late deaths occurred in 29 patients (1.8 percent), 10 months to 10.4 years after the operation. The most common cause of late death was cancer, including colon and rectal carcinoma (10 patients), hematologic malignancies (4 patients), cholangiocarcinoma (3 patients), and germ-cell carcinoma (1 patient). Four patients died from unrelated sepsis, two died following myocardial infarction, two patients died from complications of subsequent orthopedic surgery, and one patient died of cirrhosis. Two additional patients committed suicide. No late deaths were directly attributable to the IPAA procedure. CONCLUSIONS: Proctocolectomy with IPAA is a safe procedure. Operative mortality is low, and late deaths are related to carcinogenic and extracolonic manifestations of underlying or unrelated coexisting diseases and events.

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BACKGROUND: Colorectal cancer is one of the most common cancers in the world, with overall mortality exceeding 40% even with treatment. Effective efforts for screening and prevention are most likely to succeed in patient groups identified as high risk for colorectal cancer, most notably the hereditary intestinal polyposis syndromes. In these syndromes, benign polyps develop throughout the intestinal tract prior to the development of colorectal cancer, marking the patient and associated family for precancer diagnosis followed by either close surveillance or preventive treatment. PURPOSE: This review article was undertaken to discuss the most recent developments in the knowledge of hereditary intestinal polyposis syndromes, emphasizing the clinical approach to diagnosis and treatment relative to preventing the development of cancer. RESULTS: The most common of the hereditary polyposis syndromes is familial adenomatous polyposis (FAP), which is characterized by the development of hundreds to thousands of adenomatous polyps in the colon followed at an early age by colorectal cancer. Colorectal cancer can be prevented in this autosomal dominant condition by prophylactic colectomy, though a risk for other tumors, including periampullary cancers, remains throughout life. Variant of FAP associated with fewer and smaller polyps (hereditary flat adenoma syndrome), or even CNS tumors (Turcot's syndrome) also carry this high risk of colorectal cancer. Hereditary hamartomatous polyposis syndromes such as juvenile polyposis and Peutz-Jeghers syndrome (also autosomal dominant) are characterized by less frequent polyps. Though these are generally benign polyps, they are also associated with a significant risk of colorectal and other cancers. Other polyposis syndromes, including neurofibromatosis and Cowden's disease, do not carry this increased risk of colorectal cancer, and therefore affect different treatment strategies. Analysis of genetic factors responsible for these and other hereditary syndromes with predisposition to colorectal cancer has not only contributed to our molecular understanding of colorectal cancer, but opened the door to DNA testing and treatment strategies for these diseases. CONCLUSIONS: The treatment advances that are discussed and careful screening in appropriate families will effectively reduce the risk of death from colorectal cancer.

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Bispecific antibody (BsAb) with specificity for tumor cell surface antigen and the CD3 molecule on T cells can redirect activated T cells to lyse tumor cells. Since the ex vivo expansion and activation of T cells is impractical and ineffective for treating established tumors, we tested whether the immune stimulant beta glucan could in situ-activate T cells, which could secondarily be retargeted with BsAbs to lyse tumor cells. To test for tumor neutralization, C3H/HeN mice were injected i.v. with Cl-62 melanoma cells and immediately treated with i.p. beta glucan and/or anti-CD3 (500A2) x anti-p97 (96.5) F(ab')2 BsAb i.v. Pulmonary metastases were counted 14 days later. To test for tumor rejection and survival in a solid tumor model, mice were injected s.c. and i.p. with Cl-62 cells and 7 days later administered beta glucan i.p. and/or F(ab')2 BsAb i.v. In the neutralization model, there was a significant reduction in the number of metastases in the beta glucan + BsAb group, as compared with controls, and with beta glucan alone. In the established tumor model, beta glucan + BsAb reduced the incidence of s.c. tumors as compared with control, with BsAb alone and with beta glucan alone. It also prolonged survival of tumor-bearing mice compared with control, BsAb alone and beta glucan alone. We conclude that T cells can be activated in vivo by beta glucan and retargeted with F(ab')2 BsAb.

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The role of endothelial cell adhesion molecule expression in the immune response to tumors is unknown. We have investigated the expression of murine lymphocyte activation antigen (MALA-2), the murine equivalent of intercellular adhesion molecule-1 (ICAM-1), in blood vessels of normal murine tissues and in melanoma tumors and evaluated the relationship between MALA-2 expression and lymphocyte trafficking in vivo. C3H/HeN mice were injected both i.p. and s.c. with a clone of K-1735 syngeneic melanoma cells. Day 11 tumor-bearing mice were killed and vascular expression of MALA-2 was quantified using immunohistochemistry. MALA-2 expression was high in lung, liver and spleen and low in lymph node, small bowel, muscle and tumor. Systemic administration of either recombinant tumor necrosis factor alpha (rTNF alpha) or recombinant interleukin-1 alpha (rIL-1 alpha) over 2 days prior to organ harvest resulted in an increase in the number of tumor vessels expressing MALA-2, with no change in MALA-2 expression in other tissues. In vivo lymphocyte trafficking was evaluated using cultured, activated splenocytes radiolabeled with 111In. 111In-labeled splenocyte distribution correlated closely with MALA-2 expression, with high localization to spleen, liver and lung and poor localization to lymph node, small bowel, muscle and tumor. Systemic administration of rTNF alpha, but not rIL-1 alpha, resulted in a significant increase in 111In-labeled splenocyte distribution to tumor, but neither rTNF alpha nor IL-1 alpha altered 111In-labeled splenocyte distribution to normal organs. Our data demonstrate the in vivo pattern of vascular MALA-2 expression in normal murine tissues and tumors and suggest that the expression of MALA-2 can be preferentially enhanced in tumors by systemic administration of cytokines. Lymphocyte distribution in vivo correlates closely with the pattern of MALA-2 expression, and these data support the conclusion that MALA-2 plays an important role in the regulation of lymphocyte trafficking.

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OBJECTIVE: To present a large initial series of patients who underwent laparoscopic-assisted segmental colectomy and to assess the feasibility and safety of this procedure. DESIGN: We summarized the clinical outcome data for 122 Mayo Clinic patients selected for laparoscopic-assisted resection of the right, left, or sigmoid colon between 1991 and 1993. MATERIAL AND METHODS: Preexisting factors (such as obesity and prior abdominal operations), indications for surgical treatment, and intraoperative and postoperative complications were analyzed statistically in two groups of patients--those in whom the laparoscopic procedure was completed and those in whom conversion to an open surgical technique was necessary. RESULTS: Laparoscopic-assisted colectomy was successfully completed for a variety of colonic pathologic conditions, including polyps, cancer, and diverticulitis. No operative deaths occurred in this series, and the overall complication rate was low (11%). Patients in whom laparoscopic-assisted colectomy was completed had a more rapid return of bowel function and a briefer hospital stay than did those who required conversion to the traditional open surgical technique. Neither obesity nor previous abdominal surgical procedures precluded successful laparoscopic-assisted colectomy, although the conversion rate to open colectomy was 75% in patients whose weight exceeded 90 kg. CONCLUSION: These findings indicate that laparoscopic-assisted segmental colectomy is safe and feasible, and the procedure may offer patient-related advantages. Oncologic concerns, including recent reports of trocar site recurrences, suggest a cautious approach to its application for resection of colonic cancer.

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T-cell antitumor activities are limited by the requirement of two specific major histocompatibility complex restricted steps, T-helper cell activation and cytotoxic T-lymphocyte targeting. The aim of this study was to investigate whether bypassing these major histocompatibility complex restricted steps using nonspecific in vivo activation of T-cells with staphylococcal enterotoxin-B (SE-B) and retargeting with antitumor x anti-CD3 bifunctional antibody (BFA) could provide an effective antitumor response. C3H/HeN mice were injected i.v. with CL-62 melanoma cells, which express the human melanoma antigen p97, and were treated 10 min later with SE-B and/or anti-CD3 (500A2) x anti-p97 (96.5) BFA. Pulmonary metastases were counted 14 days following injections. SE-B alone induced a dose-dependent activation of T-cells as measured by increased interleukin-2 receptor expression and enhanced proliferative responses. SE-B doses greater than 10 micrograms significantly reduced the number of pulmonary metastases versus control (P < 0.01). Combined treatment with SE-B (50 micrograms) and BFA (5 to 50 micrograms) significantly decreased pulmonary metastases compared to treatment with SE-B alone (P < 0.05). Similar reductions in metastases were observed with the F(ab')2 BFA but not with the unconjugated antitumor component of the BFA. Combined treatments with SE-B plus BFA accomplished better tumor neutralization than adoptively transferred in vitro activated splenocytes (4 x 10(7)) retargeted with BFA (5-100 micrograms; P < 0.05). These studies demonstrate that T-cells can be activated in vivo by SE-B and retargeted with small doses of BFA. In this immunocompetent syngeneic pulmonary metastasis model, SE-B plus BFA provided a dramatic antitumor response.

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Antitumor x anti-CD3 bifunctional antibodies (BFAs) affect tumor cell lysis by activating and physically linking T-cells and tumor cells. Since tumor target antigen expression does not correlate with susceptibility to BFA-mediated tumor cytotoxicity, we investigated the role of cell adhesion molecules as accessory molecules. In 3 human colon tumor cell lines (LS174T, WIDR, and COLO205), recombinant interferon-gamma (rIFN-gamma) consistently increased BFA-mediated tumor cell lysis by cultured peripheral blood lymphocytes and consistently increased tumor cell expression of intercellular adhesion molecule-1 (ICAM-1). Using cell conjugation assays, we demonstrated that ICAM-1 and lymphocyte function-associated antigen-1 (LFA-1) interactions were important for effector-to-target cell conjugate formation and demonstrated that tumor cell pretreatment with rIFN-gamma enhanced cell conjugate formation. Whereas anti-LFA-1 blocked all BFA-mediated tumor lysis and conjugate formation, anti-ICAM-1 blocked only the enhancing effects of rIFN-gamma for both cytolysis and conjugate formation. Although BFAs were shown to provide effector-to-target cell bridging, LFA-1 was found to be a common critical element required for BFA-mediated cell conjugation and lysis. ICAM-1, which was augmented by rIFN-gamma, appears to be only one of several ligands interacting with LFA-1. These results provide one explanation as to why high expression of tumor-associated antigen alone does not predict the susceptibility to BFA-mediated lysis and provides further support for the concept of combined modality immune therapies.

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A prospective study was performed to determine the incidence of colorectal neoplasia and inflammatory bowel disease in patients with benign anorectal disease. Over a three-year period, 102 consecutive patients who presented with hemorrhoids, fissure, fistula-in-ano, anorectal abscess, and anal condylomata and who did not have gastrointestinal symptoms underwent colonoscopy. The mean age of all patients was 53.5 years; males out-numbered females 1.6:1. No patient was found to have inflammatory bowel disease. Ten of 102 (9.8 percent) were found to have a neoplastic lesion (nine adenomas and one adenocarcinoma). Patients found to have a neoplastic lesion tended to be older (61 years vs. 52.7 years; P = 0.06). Neoplasia was found in 4 of 21 (19 percent) with a family history of colorectal cancer and in 6 of 81 (7.4 percent) without a family history (P = 0.24). Patients presenting with outlet-type bleeding were not found to have a higher detection of neoplasia. The specific type of anorectal disease present was not associated with an increased risk for colorectal neoplasia. Our study suggests that benign anorectal disease and colorectal neoplasia may coexist. Anorectal disease is not predictive of neoplasia. The decision to perform colonoscopy should be based on age, gastrointestinal symptoms, and other risk factors.

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A possible contribution to cell toxicity in the diabetic lens due to early ATP loss is not well characterized prior to the appearance of vacuoles in the lens. Changes in lens ATP levels at longer periods of hyperglycaemia (6-8 weeks) have been reported. We used [31P]NMR analysis of lens extracts at three time periods, comparing diabetic to concurrent control groups at 1, 2 and 4 weeks of hyperglycaemia. With this design, significant alterations (> 10%) in the ATP/ADP ratio can be monitored. NMR analysis revealed a decreased ATP/ADP ratio at all time periods, averaging a 38% decrease. Luminescent determination of ATP levels indicates that this decrease is mainly caused by a decrease of 25% in ATP concentration. The early loss of GSH was large and not accompanied by an appearance of GSSG, as monitored by HPLC electrochemical detection. A 1-week experiment with animals receiving daily insulin treatment was carried out to control for effects of STZ on the lens. This treatment resulted in normal lens GSH levels and a near normal [31P]NMR profile.

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Over the past decade, flow cytometric DNA analysis has been employed by a number of investigators in an attempt to further define patient prognosis beyond classic pathologic staging. The results of these studies taken independently have been confusing; however, their cumulative effect suggests that flow cytometry is a useful prognostic indicator and can be used to further delineate prognosis within a specific pathologic stage. DNA nondiploid tumors are more likely to recur than diploid tumors, and patients with DNA nondiploid tumors have a poorer five-year survival than patients with DNA diploid tumors. There appears to be a weak relationship between advanced pathologic stage and DNA aneuploid tumors, although there is no clear and consistent relationship between tumor ploidy and histology. Therefore, all patients with colorectal tumors should undergo DNA ploidy analysis. Patients with DNA nondiploid tumors should be treated for biologically more aggressive disease independent of other prognostic variables. Ploidy status should be employed as a variable by which to randomize patients to both primary treatment schemes and adjuvant therapies in clinical trials.

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The prevention of anesthetic mishaps during endoscopic procedures is of great importance to physicians in training. With the large number of such procedures performed each year, even infrequent adverse anesthetic reactions may result in a significant number of problems. To establish the safety and efficacy of an anesthetic regimen using intravenous meperidine and diazepam, all endoscopic procedures performed at one teaching institution in a 4-month period were retrospectively analyzed with regard to: (1) type and dosage of sedation/anesthesia, (2) endoscopic procedure involved, (3) effect of any underlying disease state, (4) side effects, (5) endoscopic complications, and (6) overall patient acceptance. A total of 716 patients underwent 913 endoscopic procedures with 876 separate anesthetic/intravenous sedations. General anesthesia was utilized in 44% of the 155 pediatric procedures. In the adult patients, intravenous sedation was administered by a physician-in-training under supervision except in 9% of cases (66 patients) when intravenous sedation utilizing alternative agents was given by the anesthesia department. The dose of sedation used (per body weight) declined with increasing age in the pediatric group (0-19 years). The adult dose remained constant for the next eight decades of life (meperidine 0.76 +/- 0.33 mg/kg: diazepam 0.12 +/- 0.08 mg/kg). In the adult group, 758 procedures were performed: 371 patients underwent esophago-gastroduodenoscopy, 258 colonoscopy, 36 endoscopic retrograde cholangiopancreatography, 40 flexible sigmoidoscopy, and 51 percutaneous endoscopic gastrostomy. Anesthetic-related complications (transient apnea and itching), were noted in two patients, and naloxone was utilized to reverse oversedation in a further 17 (2.56%).(ABSTRACT TRUNCATED AT 250 WORDS)

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The increasing safety of cardiac surgery has led to the frequent referral of octogenarians for operation. Between 1980 and 1989, we reviewed our experience with 103 octogenarians (59 male, 44 female; mean age 82 years) to determine the surgical risk factors and outcome in the elderly population. There were 71 coronary bypasses (CABG), 11 aortic valve replacements (AVR), 11 AVR-CABG, 4 mitral valve replacements (MVR), 3 MVR-CABG and 3 AVR-MVR-CABG. Seventeen patients died during hospitalization (16.5%) including 9 CABG (13%); 1 AVR (9%), 2 AVR-CABG (18%), 2 MVR (50%), 1 MVR-CABG (33%) and 2 AVR-MVR-CABG (67%). Statistical analysis of 22 perioperative variables suggested that a preoperative intraaortic balloon, a history of congestive heart failure, mitral valve replacement, urgent operation, need for preoperative inotropic support and the number of anastomoses performed were significant or marginally significant (P less than 0.15) univariate predictors of operative mortality. Multivariate analysis revealed that the need for a preoperative intraaortic balloon (F = 13.1), history of congestive heart failure (F = 6.8), and MVR (F = 6.7) were significant (P less than 0.001) independent predictors of mortality. Postoperative complications included arrhythmias in 36 patients (35%), respiratory insufficiency in 11 (11%), reversible neurological deficit in 15 (14%), and a permanent neurological deficit in 6 patients (6%). Actuarial survival was 90% and 82% at 1 and 2 years, respectively. Seven of 86 (8%) long term survivors sustained a stroke in the follow-up interval. The mean follow-up of survivors was 23 +/- 19 months with a mean improvement in NYHA class of 1.4 (P less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

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Additions of micromolar concentrations of hematin to washed rat pulmonary microsomal preparations resulted in marked (5-7-fold) increases in the NADPH-dependent generation of phenolic metabolites of benzo[a]pyrene (BaP). 9-Hydroxy-BaP was identified as the major reaction product. Additions of pulmonary cytosolic fractions to microsomes produced no measurable effect but cytosol and hematin added together elicited 25-30-fold increases in total phenolic products. Cytosolic fractions from other tissues, including rat kidneys and perfused rat livers, were also highly effective in enhancing the hematin-mediated increases in monooxygenase activity. However, cytosol from human placental tissues was only minimally effective when either pulmonary or placental microsomes were utilized as enzyme source. Superoxide dismutase and catalase (alone or in combination) had no measurable effect on hematin-mediated increases. Horseradish peroxidase effectively inhibited the hematin-dependent reactions but hematin-independent reactions were inhibited with equal effectiveness. Carbon monoxide profoundly inhibited all hematin-mediated increases in metabolite formation. The activating cytosolic component was non-dialyzable, inactivated by trypsin and heat, and eluted in the void volume from Sephadex G-150 columns. This suggested that the cytosolic factor(s) responsible for the increased hematin-dependent oxidation was a protein(s) with a high molecular weight or perhaps an aggregate or oligomer of proteinaceous material. HPLC profiles indicated a major effect on the generation of phenolics; quinones were also increased but only minimal increases in diols were observed. Results were consistent with the hypothesis that hematin-mediated increases in pulmonary monooxygenase activity result from an increased association of a small pool of pulmonary P-450-apoprotein(s) with the hematin prosthetic group to result in increased levels of an unidentified holocytochrome(s) with a relatively high substrate turnover number. The current data suggest a quaternary interaction among P-450 apoprotein(s), heme prosthetic group, reaction products (particularly 3-hydroxy-BaP) and a cytosolic protein(s). We postulate that the mechanism of action of the cytosolic factor is to facilitate the interaction of hematin with the apocytochrome.

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Reaction of rat liver cadmium-metallothionein-II(Cd-MT-II) with p-hydroxymercuribenzoate(pHOHgBzO-) causes displacement of bound Cd. When pHOHgBzO- -induced displacement of 109Cd is observed after dialysis of the reaction mixture, the stoichiometry is consistent with stepwise displacement of tetraco-ordinate Cd atoms by non-random entry of reagent into the polynuclear clusters. 113Cd n.m.r. allows direct observation of the effects on bound Cd of stepwise titration of 113Cd-MT-II with pHOHgBzO-. The first equivalent reduces all resonances approximately equally. Subsequently differential reactivity of the protein thiolates towards the reagent gives rise to differential decreases in the 113Cd signal intensities. Resonances previously attributed to a three-metal cluster are lost before those arising from the four-metal cluster. These results are interpreted in terms of current models of the MT structure. They are distinct from the results of reaction of MT with 5,5'-dithiobis-(2-nitrobenzoic acid), which distinguishes between only two classes of thiolates, terminal and bridging. Such different patterns of reactivity of the protein thiolates may underlie a biological activity of this protein.

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The effects of several mono-, di-, and trithiols (400 mumol/kg) in mobilizing Cd from metallothionein were studied in rats 24 h after a single injection of 109CdCl2 (1 mg kg/Cd). BAL (2,3-dimercaptopropanol) and propanetrithiol (1,2,3-trimercaptopropane) were the most effective mercaptans in increasing the biliary excretion of Cd (3.1 and 5.5% of administered dose, respectively, compared to 0.04% in control injected with propylene glycol) in in situ experiments with a significant decrease in hepatic Cd. Propane-1-thiol was inactive and propanetrithiol was the most toxic of the compounds studied. A number of propane dithiols having sulfhydryl groups at different carbon positions with and without substituents (OH or SO-3) and dimercaptoethane were also tested for effectiveness in removing Cd. All the lipophilic compounds with two adjacent sulhydryl groups were effective in increasing the biliary excretion of Cd, but were less effective than BAL and propanetrithiol. The major form of Cd in liver cytosols of rats pretreated with CdCl2 and injected with mercaptans was metallothionein. A small amount of Cd in liver cytosol and a major portion of biliary Cd in propanetrithiol-injected rats were bound to high-molecular-weight proteins when fractionated on Sephadex G-75 columns. On the other hand, after injection of BAL, most of the Cd in the bile was associated with a fraction of molecular weight 10,000. Even though Cd was present mainly as metallothionein in livers of Cd-pretreated rats, the biliary forms of Cd after injection of BAL and propanetrithiol were different. Similar results were obtained when Cd was added in vitro to bile samples collected from control rats that were injected with these chelating agents alone. However, the Sephadex G-75 elution profile of Cd-BAL and Cd-propanetrithiol after direct addition to control rat bile showed Cd complexes of identical molecular weight (less than 6000). These results suggested that the Cd-binding ligands present in bile after injection of BAL and propanetrithiol were different from and had a higher molecular weight than the complexes in vitro with Cd and the respective chelating agents.

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