Carcinoembryonic antigen induces cytokine expression in Kuppfer cells: implications for hepatic metastasis from colorectal cancer.

The mechanism by which carcinoembryonic antigen (CEA) causes enhancement of hepatic metastasis from colorectal cancer is not defined. We hypothesize that binding of CEA to an 80-kDa Kupffer cell receptor by the peptide sequence Pro-Glu-Leu-Pro-Lys (PELPK) induces cytokine production in the hepatic microenvironment, which then impacts on the formation of hepatic metastasis from colorectal cancer. We have, therefore, isolated Kupffer cells and treated them in vitro with CEA, its gene family member nonspecific cross-reacting antigen, PELPK-albumin conjugate, and lipopolysaccharide as a positive control. Spent media was examined for the content of cytokines interleukin (IL) 1alpha, IL-1beta, IL-6, and tumor necrosis factor alpha using the ELISA. Simultaneously, mRNA was extracted from the same cells and amplified using reverse transcription-PCR to evaluate the induction of specific cytokine transcripts. As expected, lipopolysaccharide stimulated cytokine production. CEA, nonspecific cross-reacting antigen, and PELPK-albumin induced secretion of all of the cytokines tested; the response was higher in general with PELPK-albumin. The levels of cytokine mRNA showed a similar profile. These responses were not seen when a similar but irrelevant peptide conjugate PELGK-albumin was used. These results demonstrate that binding of the peptide PELPK to the 80-kDa receptor results in the release of a series of cytokines that have the potential to activate hepatic sinusoidal endothelium. This may explain CEA-induced enhancement of experimental hepatic metastasis.

Purification and analysis of an 80-kDa carcinoembryonic antigen-binding protein from Kupffer cells.

The receptor-mediated interaction of Kupffer cells with carcinoembryonic antigen (CEA) has led to the identification of an 80-kDa CEA-binding Kupffer cell protein. This study is aimed at the isolation and analyses of this protein from rat Kupffer cells. The binding protein was purified using a combination of gel filtration, preparative polyacrylamide gel electrophoresis (PPAGE), and affinity chromatography using a CEA-Sepharose column. Fractions obtained from the gel filtration produced two major and few minor peaks with CEA-binding activity. Maximum reactivity was detected in the first major peak. The first major peak protein was partially precipitated following fractionation with 30% loss of activity in the precipitate. Fractions with CEA-binding activity were pooled and separated on the basis of molecular weight (MW) in PPAGE. The fractions between MW 70 and 90 kDa were pooled and affinity purified using CEA-Sepharose affinity chromatography. The purity of the 80-kDa protein was demonstrated by a single protein band on SDS-polyacrylamide gel. The protein was further identified by an anti-80-kDa binding protein antibody in Western blot analysis. The pI of the 80-kDa protein is 4.95. Amino acid analysis demonstrated no histidine; higher percentages of glutamine (13.3%), leucine (11.2%), asparagine and alanine (10.4%), and lysine (9.2%) were observed. Protein microsequencing revealed two unique sequences, one with 16 amino acids and the other with 11 amino acids. The 16-amino-acid sequence has less than 50% homology with a large sample of unrelated proteins, whereas the sequence containing 11 amino acids has 60-70% homology with the alpha chain of collagen from a variety of species but no significant homology with other known proteins, suggesting the presence of collagen-like domains in the 80-kDa receptor.

Exercise testing and training of patients with malignant ventricular arrhythmias.

Malignant ventricular arrhythmias account for a significant number of deaths annually. Improved therapies have increased survival for this patient population. These patients, however, remain at high risk due to their history of sudden cardiac death and the preponderance of poor ventricular function in this population. This high risk status may have excluded them from participation in cardiac rehabilitation programs or from undergoing exercise testing in some medical centers. Although the risk of complications, especially arrhythmias, is quite real, the experience in some centers has indicated that it is manageable. This experience and some guidelines for exercise testing and training of patients with malignant ventricular arrhythmias are reviewed in this paper.

Spirometry as a preoperative screening test in morbidly obese patients.

We performed spirometry on 114 morbidly obese patients considered for gastric bypass surgery to assess its efficacy as a preoperative screening test. One hundred eight subjects underwent surgery, and 61 patients returned for repeat spirometry 1 year later. The average preoperative forced vital capacity (FVC), forced expiratory volume in 1 second (FEV1), and mid flow (FEF25-75%) were 100% of predicted. Spirometry identified no more of our obese subjects as abnormal than would have been identified in a group of healthy, nonobese individuals. Each surgical case was reviewed. An abnormal preoperative spirogram did not identify the patients who experienced postoperative complications. Weight loss was associated with very small increases in FVC (300 ml) and FEV1 (245 ml). Preoperative spirometric testing is not indicated in morbidly obese patients with no other identifiable risk factors for postoperative respiratory complications.

Membrane association of soluble protein activators of rat liver adenylate cyclase. Evidence for distinctness from the guanine nucleotide-binding stimulating protein (Ns).

Sonication of a crude rat liver membrane preparation and centrifugation at 100,000 X g yielded a supernatant which activated basal and hormone-sensitive adenylate cyclases [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1]. The membrane origin of the stimulatory activity was confirmed by the use of lactate dehydrogenase as a marker for contamination by cytosol. The solubility of the activating factors was verified by their passage through 0.05 micron diameter pores of Millipore filters. The membrane-derived activators were nondialyzable and destroyed by heat and trypsin in the same manner as adenylate cyclase activators detectable in cytosol. Stimulation by factors from membranes and cytosol was not additive. The amount of the activators which could be freed from membranes by sonication was 12-15% of that contained in cytosol previously separated from the membranes. Soluble activators from the two sources had limited ability to restore adenylate cyclase activity to membranes from the cyclone of S49 mouse lymphoma cells which are deficient in the enzyme's guanine nucleotide-binding stimulatory protein, Ns. Cytosol did not contain a substrate for ADP-ribosylation by cholera toxin that corresponded electrophoretically to Ns. Furthermore, purified Ns did not affect adenylate cyclase activity in preparations stimulated by the soluble activators. These findings suggest that the activating factors found in cytosol may be released from membranes during tissue homogenization. Because these protein activators can be obtained from membranes without use of detergents and can neither substitute for nor be substituted for by Ns in functional assays, they are distinct from Ns.

Changes in serum lipids after gastric bypass surgery. Lack of a relationship to weight loss.

Serum lipid levels were measured in 65 obese patients before and a mean of 13.4 +/- 0.2 months after gastric bypass surgery performed to facilitate weight reduction. The 58 women studied lost an average of 39.8 +/- 1.2 kg thereby reducing their mean body mass index from 41.9 +/- 0.7 to 26.8 +/- 0.6. Postoperatively, total cholesterol, LDL cholesterol, and triglyceride levels were significantly lower and HDL cholesterol levels significantly higher than preoperative values (for each of the four comparisons P less than 0.0001). Whereas before surgery 83 percent of the women had levels of total, LDL, or HDL cholesterol indicative of an increased risk of coronary heart disease, after surgery only 24 percent did so. Preoperative cholesterol abnormalities in the seven men were not as severe as were those in the women, and the men's postoperative cholesterol changes were not as marked. Correlations of the difference between pre- and post-operative weight and the difference between pre- and post-operative lipid levels showed that weight loss accounted at most for just 8 percent of the simultaneous lipid change. The data indicate that gastric bypass surgery for morbid obesity produces a favorable change in serum lipid profiles and suggest that the change is due primarily to factors other than the associated weight reduction.

Hypoxemia after gastric bypass surgery for morbid obesity.

Fifty-six patients who underwent Roux-en-Y gastrojejunostomy for morbid obesity had arterial blood gas analysis before surgery and during the first five postoperative days. Preoperatively, seven subjects were hypoxemic and three were hypercapneic. Twenty-four hours after gastric bypass, 75% of the patients had an arterial oxygen pressure (Pao2) less than 60 mm Hg. Compared with preoperative measurements, blood gas values on the first postoperative day showed a 13.7-mm Hg decrease in the mean arterial Pao2 and a 5.0-mm Hg increase in the mean arterial carbon dioxide pressure. Arterial Pao2 determinations on the third, fourth, and fifth postoperative days returned toward, but remained significantly less than, paired values obtained before surgery. Patients with hypoxemia after gastrojejunostomy were significantly older and had significantly lower preoperative arterial Pao2 measurements than patients who were not hypoxemic after surgery. Weight, body mass index, and preoperative spirometric measurements did not distinguish between those patients who did and did not become hypoxemic postoperatively. We conclude that hypoxemia commonly follows gastric bypass for morbid obesity, and thus we recommend that all patients undergoing this procedure be treated with supplemental oxygen for at least the first three postoperative days.

Ectopic beta-adrenergic receptors coupled to adenylate cyclase in human adrenocortical carcinomas.

The adenylate cyclase of an adrenocortical carcinoma of the rat is activated not only by ACTH but also by beta-adrenergic agonists, which bind to ectopic beta-adrenergic receptors not present in normal rat adrenal cortex. Previous reports examining possible beta-adrenergic control of adenylate cyclase in human adrenocortical carcinomas failed to demonstrate beta-adrenergic receptor-linked enzyme activity. We studied six human adrenal carcinomas and normal adrenal cortex from three subjects for beta-adrenergic agonist-sensitive adenylate cyclase and beta-adrenergic binding sites. Three of the six carcinomas had adenylate cyclase responses to both ACTH and beta-agonists. Two tumors were ACTH responsive but not beta-agonist responsive; one tumor responded to beta-agonists but not to ACTH. Adenylate cyclase activity of normal adrenal cortex from three subjects was stimulated by ACTH but not by beta-agonists. In membrane preparations from three tumors with beta-agonist-sensitive adenylate cyclase, the radiolabeled beta-adrenergic antagonist [125I]pindolol bound specifically and with high affinity (Kd = 38-83 pM) to a single class of binding sites which showed saturation with ligand concentration, reversibility of binding, pharmacological specificity, and stereospecificity. Normal cortex and one tumor without beta-adrenergic agonist-sensitive adenylate cyclase had no specific binding of [125I]pindolol. These results indicate that malignant transformation of adrenal cortex in man is frequently but not invariably associated with the appearance of ectopic beta-adrenergic receptors functionally linked to adenylate cyclase. Loss of ACTH-responsive adenylate cyclase may also occur simultaneously with the development of beta-adrenergic receptor-linked adenylate cyclase.

Hypogonadism in hemochromatosis: reversal with iron depletion.

Gonadal function was evaluated in 64 persons homozygous for the HLA-linked hemochromatosis allele. Of 41 men, 10 had reduced libido or impotence and 6 had testicular atrophy. Before treatment, 5 men had below normal testosterone concentrations, 4 of whom also had low gonadotrophin levels. Four hypogonadal men were reevaluated after iron depletion treatment. In 2, 1 with primary and another with secondary hypogonadism, testosterone levels returned to normal after phlebotomy and were accompanied by a return of normal sexual function. None of 23 women with hemochromatosis had loss of libido or had a natural menopause before age 45. Our findings indicate that in some men with hereditary hemochromatosis and hypogonadism of either testicular or central origin, sexual function and sex hormone concentrations can be restored to normal after iron depletion therapy.

Systemic effects of oral contraceptives.

PIP: Reduced estrogen content has significantly decreased the risks of oral contraceptive (OC) use. However, the systemic effects of OCs, but it is unclear if this change is physiologically significant. Estrogen-mediated inhibition of cortisol levels may contribute to the impairment of glucose tolerance by OCs. Women at high risk for diabetes, older than 35, obese, with family history of diabetes, or who have had glucose intolerance during previous pregnancies should either not take OCs or take pregestin-only pills. OCs raise plasma triglyceride levels 30-50 mg per dl in users of all ages. High density lipoprotein (HDL) cholesterol is also affected, and cholesterol and triglyceride levelshould be measured before and during OC use. The risk of hepatic adenoma rises with duration of OC use; however, most adenomas diagnosed before hemorrhage have regressed with discontinuation of the contraceptive regimen. The most significant adverse effects of OC use involve the arterial and venous vascular systems. OCs appear to raise the blood pressure in nearly all women. Change in systolic pressure is consistently greater than in diastolic, suggestingthat the primary hypertensive effect of OCs is on blood volume and cardiac output. Accumulated data indicate that if OCs are not used by women older than 35 or by women who smoke or who are hypertensive, then risk of subarachnoid hemorrhage or other cerebrovascular complication is very small. The relative risk of myocardial infarction in OC users has been from 0-6 times greater than in nonusers; this may depend on other confounding risk factors. Reduction in estrogen content of OCs decreases risk accordingly. The preponderance of evidence indicates that prolonged use of OCs does not increase risk of breast disease or ovarian and endometrial cancer, and, in fact, may protect users from malignant lesions by suppressing gonadotropins and ovulation.^ieng

Anions and cations as stimulators of liver adenylate cyclase.

The effects of inorganic salts on adenylate cyclase activities were studied in rat liver homogenates, particulates, and purified membranes. Na+ salts of N-3, NO-2, S2O32-, SO42-, and non-F- halides stimulated homogenates by a maximum of 2---5-fold; half-maximal effects were seen at concentrations ranging from 90 mM (Na2S2O3) to 410 mM (NaNo2). NaIO3 stimulated about 2-fold at relatively low salt concentrations (5---20 mM). Na+ salts were stimulatory using both ATP and the ATP analog 5'-adenylyl-beta, gamma-imidodiphosphate (AMP-P(NH)P) as enzyme substrate. The time courses of stimulation by NaCl and NaN3 were linear to at least 10 min with any of the three tissue preparations. Although salt effects clearly varied with the different anions, the magnitude and dose-response of stimulation of homogenates by Cl- salts were also dependent on the accompanying alkali cation (Li+, Na+, K+, Rb+, Cs+). MgCl2 at relatively high concentrations (50 mM) enhanced NaCl-stimulated activity in homogenates only slightly at relatively low concentrations of NaCl and not at high concentrations, suggesting a common mechanism of activation by Mg2+ and Na+ salts. NaCl- and NaN3-stimulated activities were unstable in homogenates kept at 0 degree C for 4 h, falling to 35% and 50% of their respective baseline activities. The effects on homogenates and particulates of maximally stimulatory concentrations of NaCl and NaN3 were further enhanced by GTP and the GTP analog 5'-guanylyl-beta, gama-imidodiphosphate (GMP-P(NH)P).