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Congenitally jaundiced (jj) Gunn rats had a greater hepatic microsomal content of a cytochrome P-450 isoenzyme, P-450c, than did the non-jaundiced (Jj) rats. No differences in content of P-450b, P-450d and pregnenolone-16 alpha-carbonitrile-induced (PCN) P-450 were found between jj and Jj rats. This is the first demonstration of a constitutive increase in a specific cytochrome P-450 isoenzyme in association with a genetic defect.

In seeking to identify nucleating/antinucleating proteins involved in the pathogenesis of cholesterol gallstones, a major acidic protein was isolated from each of 13 samples of cholesterol gallstones. After the stones were extracted with methyl t-butyl ether to remove cholesterol, and methanol to remove bile salts and other lipids, they were demineralized with EDTA. The extracts were desalted with Sephadex-G25, and the proteins separated by PAGE. A protein was isolated, of molecular weight below 10 kD, which included firmly-bound diazo-positive yellow pigments and contained 24% acidic, but only 7% basic amino acid residues. The presence of N-acetyl glucosamine suggested that this was a glycoprotein. This protein at concentrations as low as 2 micrograms/ml, but neither human serum albumin nor its complex with bilirubin, inhibited calcium carbonate precipitation from a supersaturated solution in vitro. This protein could be precipitated from 0.15 M NaCl solution by the addition of 0.5 M calcium chloride. Considering that cholesterol gallstones contain calcium and pigment at their centers, and that small acidic proteins are important regulators in other biomineralization systems, this protein seems likely to play a role in the pathogenesis of cholesterol gallstones.

The effect of a mixture of amino acids (MAA) perfusion of the proximal jejunum on the secretory response of volume, bicarbonate, enzymes, protein, and total bile salts was studied in duodenal aspirates from 15 normal volunteers. The response was compared in the same subjects on different days with that to continuous intravenous infusion of cholecystokinin-pancreozymin (CCK), in maximally tolerated doses and to a control saline perfusion of the jejunum. A three-lumen tube was used for: perfusion of duodenal marker (phenol red), distal aspiration of duodenal samples, and jejunal perfusion of MAA or saline, 40 cm beyond the ligament of Treitz. Fifteen-minute duodenal collections were made for 1/2 hr before, 2 hr during, and 1 hr after stimulation. The results suggested that jejunal MAA perfusion releases endogenous CCK which is as effective a stimulus of pancreatic enzyme secretion as continuous intravenous CCK infusion. Both jejunal MAA perfusion and intravenous CCK infusion also stimulated volume and bicarbonate output in duodenal aspirates. In addition, jejunal MAA perfusion in the 2nd hr produced a sustained increase in total bile salt output which is probably related to continued gallbladder contraction. We concluded that physiological concentrations of amino acids in the proximal jejunum act as an important factor in the regulation of exocrine pancreatic and biliary secretions in man.

Plasma levels of the major norepinephrine (NE) metabolite, 3-methoxy-4-hydroxyphenylglycol (MHPG), were measured in a rapidly cycling bipolar patient and her first-degree relatives. The mood state dependence and the reciprocal relationship between noradrenergic and cholinergic activity were investigated by assessing mood, thought disorder, and plasma MHPG following the infusion of physostigmine. A correlation was found between plasma MHPG and mood states, with exceedingly high levels during mania and hypomania. Levels were significantly decreased by ECT or combined lithium and chlorpromazine administration. A pathologic diurnal MHPG pattern was detected during periods of abnormal mood changes. Infusion of physostigmine led to a prompt reduction in MHPG release and a marked decline in mood-state measurements and the overall level of thought disorder. Muscarinic receptors exerting negative feedback control over the synthesis and/or release of NE may have become supersensitive as a consequence of alpha 2-adrenoceptor densensitization.

"New generation" antidepressants are generally considered to include all agents introduced in recent years which are neither tricyclic in structure nor monoamine oxidase inhibitors. They include the tetracyclics (mianserin and maprotiline), the bicyclic serotonergic compounds (fluoxetine and citalopram), and the unicyclics (bupropion), as well as the triazolobenzodiazepine derivatives (alprazolam), the triazolopyridines (trazodone) and the tetrahydroisoquinolines (nomifensine). Methodologic and economic considerations have hampered attempts to develop agents with significantly greater specificity or safety than traditional agents. Traditional agents, while lacking specificity, do have extensive records of efficacy and long-term safety and are usually less expensive than new agents. Patients not responding to traditional agents often have medical or characterologic problems that exclude them from participating in controlled studies of new agents. These problems are discussed and potential approaches to the development of new agents are presented.

We assessed the effects of in vivo phototherapy of Gunn rats on the activity of hepatic microsomal mixed-function monoxygenases and on the in vivo pharmacokinetics of [14C]HB. In experiment 1 no serial changes were seen in activities of hexobarbital hydroxylase or benzo(a)pyrene hydroxylase in hepatic microsomes isolated after 2, 4 or 7 days from homozygous jaundiced female Gunn rats exposed to continuous phototherapy or in matched Gunn rats maintained under dim light. In experiment 2 homozygous jaundiced (jj) and heterozygous nonjaundiced (Jj) Gunn rats of both sexes each received i.v. [14C]HB on 2 successive days. In random order, each was exposed on the first or second day to phototherapy for 5.5 hr, beginning 0.5 hr before the administration of HB; otherwise, each was kept under dim light. Plasma [14C]HB in arterial blood samples was separated chromatographically from its labeled metabolites, and biexponential plasma disappearance curves for [14C]HB were analyzed by a SAAM-23 computer program. Clearances in female rats were much slower. In both sexes, the total body clearance and volume of distribution of HB were decreased by 20% during phototherapy of the jj but not the Jj rats; terminal plasma half-life was unchanged. In experiment 3 direct in vitro illumination of [14C]HB did not cause photodegradation of this compound, despite the presence of albumin with or without bilirubin.

We previously reported that 27% of 92 cholecystectomized patients had pigment stones (Am J Dig Dis 19:585-590, 1974). Using standard biochemical methods, we found that cholesterol accounted for an average of 77% of the dry weight of cholesterol stones, but that unconjugated bilirubin represented a mean of only 7% of pigment stones. This quantitation of pigment stones was limited because approximately 66% of their weight was insoluble. To characterize pigment and cholesterol stone composition further, we used infrared spectroscopy--a technique requiring neither crystallinity nor solubilization--to quantitate pigment, carbonate, and cholesterol in gallstones. Other organic and inorganic components of stones were measured by standard methods. By infrared spectroscopy, two types of pigment stones were identified: carbonate-containing and noncarbonate pigment stones. Carbonate pigment stones contained significantly more calcium, carbonate, and phosphate, but less pigment than noncarbonate stones. Compared to our initial report, the total measured components of all pigment stones were increased 6-fold from 10 to 63%. Cholesterol was the major component of cholesterol stones by chemical assay or infrared spectroscopy. Among five cholesterol stones with limited solubility, 80% of the insoluble residue was identified as cholesterol by infrared spectroscopy. This study extends our knowledge of pigment stone and cholesterol stone composition by the use of quantitative infrared spectroscopy in conjunction with standard biochemical methods; furthermore, it confirms that pigment and cholesterol stones differ in composition and form by different mechanisms.

A new thin-layer chromatographic (TLC) method is described for measurement of conjugated (CB) and unconjugated (UCB) bilirubin in bile. Bile is streaked on a silica-gel plate and developed with chloroform. Bilirubin in the mobile UCB band and stationary CB band is quantitated by diazotization from the scraped gel with p-iodoaniline reagent. Both CB and UCB in bile yielded identical azopigment absorbance. There was no contamination of the UBC band with either CB or oxidative derivatives of bilirubin, and over 98 per cent added 14C-UBC migrated with the mobile band. Recoveries from TLC averaged 98.2 +/- 4.3 per cent of the applied pigment, duplicated agreed within 10 per cent, and the method accurately detected the enzymatic and alkaline hydrolysis of CB to UCB. Concentrations of UCB in nine samples of normal human gallbladder bile were found to be less than 1.9 mg./dl. and never exceeded 1.2 per cent of the total bilirubin.

The i.r. spectra of bilirubin isomers that differ in number and position of vinyl groups were examined to verify the assignment of the 988 cm−1 peak of bilirubin (991 cm−1 peak in calcium bilirubinate) to its pendant vinyl groups. There were only small changes in this peak with changes in position of vinyl groups (exo-2- and −18-vinyl versus endo-3- and −17-vinyl), but progressive loss of peaks in this region was observed when vinyl groups were reduced to ethyl groups (dihydrobilirubin and mesobilirubin). Methylvinylmaleimide, a monopyrrole derived from the outer (A and D) rings of bilirubin, has a pendant vinyl group and exhibits a prominent peak at 986 cm−1, but haematinic acid methyl ester derived from the inner (B and C) rings has no vinyl group and shows no peak near 988 cm−1. These observations verify the assignment of the 988 cm−1 peak of bilirubin to its pendant vinyl groups. This supports our previous proposal that a decrease in the peak at 985-995 cm−1 in the i.r. spectra of pigment gallstones, as compared with unconjugated bilirubin or calcium bilirubinate, indicates a consumption of vinyl groups in the process of formation of the polymer in the pigment stones.

To determine whether the frequency of chronic hepatitis B in homosexual men reflects the high rate of acute hepatitis B or an altered response to hepatitis B, 236 homosexual men were prospectively followed up after entry into the placebo group of a hepatitis B vaccine trial. Sixty-six participants (28.0%) developed hepatitis B ten to 18 months after entering the study. Only four (6.1%) of the 66 had persistence of hepatitis B beyond six months, similar to reports in nonhomosexual subjects. This study suggests that the high rate of acute hepatitis B is the major factor contributing to the high prevalence of chronic hepatitis B in homosexual men, rather than an altered host response. Clinical features of the acute illness could not be used to identify patients who developed chronic hepatitis B.

The studies reviewed here and ongoing work indicate a primary role for psychiatry in treating patients with the acquired immunodeficiency syndrome (AIDS) and human immunodeficiency virus (HIV) infection. Those patients often have neuropsychiatric disorders concurrent with the complications of physical disease. It is important to understand the diagnostic criteria for patients with HIV-related disorders to distinguish CNS disturbances caused by infections from those psychiatric problems exacerbated by the social consequences of the syndrome. Although treatment is difficult in immunocompromised patients and treatments may cause serious side effects, many of the CNS symptoms of AIDS patients can be treated in the same way as other diseases of the CNS. Psychostimulants are the most promising therapeutic agents for AIDS patients. In the treatment of psychotic episodes, high-potency neuroleptics at low dosages have been used successfully. In all therapies, careful consideration must be given to possible adverse reactions, which may be more serious in HIV-related diseases.

A randomized, crossover trial was carried out on the effect of moderate sodium reduction on red-blood-cell sodium metabolism. The participants were healthy high school students (mean age = 16 years, n = 33). Changes in sodium-lithium countertransport and intracellular sodium concentration were evaluated 24 days after a decrease in dietary sodium from approximately 110 to 40 mEq per day. Dietary sodium restriction had no significant effect on either sodium-lithium countertransport or intracellular sodium concentration.

Blood pressure screening was carried out on a university campus to identify early hypertension or high-normal BP in young adults. Compared with normotensive control subjects of a similar age, drawn from the same population, persons identified as being at the upper end of the BP distribution had significantly increased levels of sodium-lithium countertransport. This difference persisted when other potential confounding variables, eg, overweight, sex, ethnicity, sodium excretion, and age, were taken into account. A positive family history was associated with slightly higher levels of sodium-lithium countertransport, although the effect could be explained by higher present levels of BP. These data suggest that abnormalities of cation transport are present early in the course of the development of hypertension. Measurement of transport levels may provide an estimate of risk of hypertension and allow identification of susceptible persons.

Sixty-five gallstones were assessed for content of network polymer by equilibrium swelling and infrared spectroscopy. All types of pigment gallstones exhibited swelling in aqueous buffers, indicating network polymer content, but swelling was greatest in black pigment gallstones. Gallstones composed principally of cholesterol did not swell. Swelling increased with increasing pH values from 6 to 10.5, suggesting that titratable carboxyl side chains of the pigment were intact. Swelling increased in the presence of 10 mM ethylenediaminetetraacetic acid, indicating that calcium ions provided secondary crosslinks in the network. In all types of pigment gallstones, the equilibrium swelling ratio at the maximum (pH = 10.5) was correlated inversely with the proportion of intact vinyl groups in the pigment assessed by infrared spectroscopy. This suggests that pigment vinyl groups are consumed in forming the covalent crosslinks of the polymer network. A weaker direct correlation between equilibrium swelling ratio and the percentage of unmeasured residue in the gallstones suggested that this residue may also have contributed to the swelling. The high content of cross-linked network polymer in black pigment gallstones renders their therapeutic dissolution improbable by physicochemical means.

Further evidence that the bilirubin excreted by Gunn rats during phototherapy is unconjugated is presented. The unconjugated bilirubin is not carried into bile as a water-soluble complex with rapidly excreted bilirubin derivatives, nor as a result of leakage of plasma protein into bile. Phototherapy does not augment biliary excretion of bilirubin in normal nonjaundiced rats, but does increase the concentration of conjugated bilirubin in the bile of patients with alcoholic cirrhosis, although their serum bilirubin concentrations are unaffected. The mechanism for augmented excretion of unconjugated bilirubin during phototherapy remains unexplained.

Pruritus associated with hepatic cholestasis may cause significant morbidity. Current evidence indicates that bile acids may not be the pruritogenic factor. Rather, the factor may be some other cholephilic anion or anions that bind to cholestyramine resin and are photolabile. Effective treatment modalities used currently include cholestyramine resin and phenobarbital. Phototherapy is a promising modality in which the mechanism and most effective wavelengths remain to be elucidated. Analysis of the possible role of non-steady state bile acid compartmentalization in the pathogenesis of pruritus is presented.

New information is summarized, indicating that configurational photoisomerization of bilirubin at the 5 and 15 carbon bridges is the major mechanism of bilirubin photocatabolism in vivo, and that singlet oxygen photooxidation plays only a minor role. The literature is reviewed concerning potentially damaging photodynamic reactions that are observed in vitro with vitamins, proteins, lipids, and nucleic acids, and their possible relationships to the limited number of toxic side-effects that have been detected with clinical phototherapy of neonatal jaundice. Secondary toxic effects, mediated by bilirubin photoderivatives or by retina-neuroendocrine pathways are also considered. Areas requiring further investigations are delineated.

A 19-year-old woman with extensive, persistent chronic graft versus host disease (GVHD), following an HLA-identical bone marrow graft for acute leukemia, developed rapidly progressive airflow obstruction 140 days post-transplantation (PT) and presented clinically with persistent cough, inspiratory rales, bronchospasm and exertional dyspnea. Pulmonary function tests (PFT) showed rapidly evolving severe airflow obstruction and hypoxemia without restrictive ventilatory defect. Open lung biopsy on the 204th day PT confirmed focal bronchiolitis obliterans. On the 381st day PT, she remained clinically stable. Chest x-ray film showed mild overinflation, but was otherwise unremarkable. PFT's continued to show very severe airflow obstruction without restrictive ventilatory defect. The etiology of the obliterative bronchiolitis might be explained on the basis of a direct immunologic reaction mediated by GVHD or possibly a joint viral-GVHD interaction. Awareness and further detailed documentation and analysis of this unusual respiratory syndrome associated with marrow transplant recipients may help clarify the role of GVHD in the development of lung disease in recipients of marrow grafts.

Phototherapy has been used to treat neonatal jaundice, but little assessment has been made of possible beneficial effects on adult liver disease. Effects of phototherapy on bile acid turnover, biliary lipid concentration, liver function tests, and bromosulfophthalein (BSP) kinetics were studied in 8 alcoholic cirrhotics. Phototherapy initially increased biliary specific activity of both primary bile acids and then produced an acceleration of cholic and chenodeoxycholic acid decay curves. Pruritus was relieved in the 3 patients who had this symptom. The proposed mechanism for these changes is mobilization of bile acids from an expanded cutaneous bile acid pool with augmented bile acid excretion. No significant change in serum liver function tests or BSP plasma disappearance curves was seen. Phototherapy causes little improvement in intrinsic liver function, but produces specific changes in bile acid metabolism; these changes may be related to effects of light on a cutaneous bile acid pool.

Pigment gallstones are defined as any dark brown-to-black stone, consisting of calcium salts of bilirubin, phosphate, carbonate and other anions, and can be separated into carbonate- and noncarbonate-containing groups. Pigment stones predominate in the rural Orient, in cirrhosis, and in elderly United States patients undergoing cholecystectomy. Clinical associations include bile duct obstruction, stasis, and possibly hemolysis. Of pigment stones, 50% are radioopaque and account for two-thirds of all opaque stones. The concentrations of bile salts, phospholipids,, cholesterol, and total bilirubin in bile are similar to normal levels, but the concentration of unconjugated bilirubin is increased in the bile of some patients. Increased unconjugated bilirubin in bile may be caused by increased hydrolysis of excreted conjugated bilirubin. Unconjugated bilirubin is solubilized by bile salts, but the interaction is primarily nonmicellar. Ionized calcium and pH are important determinants of solubility. Sulfated glycoproteins, excreted in increased amounts in patients with cholelithiasis, may be the site of pigment stone precipitation because these compounds bind calcium salts tightly. E coli is frequently cultured from pigment stones in Japan but not in the United States; thus, bacterial beta-glucuronidase may be important in stone formation in Japan but probably not in the West. Stasis leads to increased calcium secretion and to increases in the concentration of sparingly soluble compounds that may then precipitate. Incomplete emptying of the gallbladder may result in the same concentration process. Unsaturated fats and chronic vagal stimulation cause pigment stone formation in animals. At present, surgery is the only treatment for pigment lithiasis.

2507 homosexual men who were seronegative for human immunodeficiency virus (HIV) at enrollment were followed for six months to elucidate risk factors for seroconversion to HIV. 95 (3.8%) seroconverted. Of men who did not engage in receptive anal intercourse within six months before baseline and in the six-month follow-up period, only 0.5% (3/646) seroconverted to HIV. By contrast, of men who engaged in receptive anal intercourse with two or more partners during each of these successive six-month intervals, 10.6% (58/548) seroconverted. No HIV seroconversions occurred in 220 homosexual men who did not practise receptive or insertive anal intercourse within twelve months before the follow-up visit. On multivariate analysis receptive anal intercourse was the only significant risk factor for seroconversion to HIV, the risk ratio increasing from 3-fold for one partner to 18-fold for five or more partners. Furthermore, data for the two successive six-month periods show that men who reduced or stopped the practice of receptive anal intercourse significantly lowered their risk of seroconversion to 3.2% and 1.8%, respectively. Receptive anal intercourse accounted for nearly all new HIV infections among the homosexual men enrolled in this study, and the hazards of this practice need to be emphasised in community educational projects.

A criticial evaluation was made of the ethyl anthranilate diazo and two solvent-partition methods for the determination of conjugated and unconjugated bilirubin in human and rat bile. The ethyl anthranilate diazo reagent, which reacts only with conjugated bilirubin in serum, also diazotized a variable proportion of unconjugated bilirubin in bile and thus overestimated the concentration of monoconjugates. With the Weber-Schalm and modified Folsch solvent-partition methods applied to human or rat bile, 4–9% of added 14C-labelled unconjugated bilirubin partitioned with the conjugated bilirubin in the upper phase, and 4–9% of added 14C-labelled conjugated bilirubin partitioned into the lower phase. With dog bile, the spill-over of 14C-labelled bilirubin into the lower phase was 9–11%. Analysis of azopigments from the Weber-Schalm partition confirmed that over two-thirds of the bilirubin in the lower phase represents monoconjugates, principally the less-polar monoxylosides and monoglucosides. These solvent-partition methods thus overestimate the concentration of unconjugated bilirubin in bile.

The feline leukemia virus subgroup C receptor (FLVCR) is a heme export protein that is required for proerythroblast survival and facilitates macrophage heme iron recycling. However, its mechanism of heme export and substrate specificity are uncharacterized. Using [(55)Fe]heme and the fluorescent heme analog zinc mesoporphyrin, we investigated whether export by FLVCR depends on the availability and avidity of extracellular heme-binding proteins. Export was 100-fold more efficient when the medium contained hemopexin (K(d) < 1 pm) compared with albumin (K(d) = 5 nm) at the same concentration and was not detectable when the medium lacked heme-binding proteins. Besides heme, FLVCR could export other cyclic planar porphyrins, such as protoporphyrin IX and coproporphyrin. However, FLVCR has a narrow substrate range because unconjugated bilirubin, the primary breakdown product of heme, was not transported. As neither protoporphyrin IX nor coproporphyrin export improved with extracellular hemopexin (versus albumin), our observations further suggest that hemopexin, an abundant protein with a serum concentration (6.7-25 mum) equivalent to that of the iron transport protein transferrin (22-31 mum), by accepting heme from FLVCR and targeting it to the liver, might regulate macrophage heme export and heme iron recycling in vivo. Final studies show that hemopexin directly interacts with FLVCR, which also helps explain why FLVCR, in contrast to some major facilitator superfamily members, does not function as a bidirectional gradient-dependent transporter. Together, these data argue that hemopexin has a role in assuring systemic iron balance during homeostasis in addition to its established role as a scavenger during internal bleeding or hemolysis.

Background The unbound, free concentration (Bf) of unconjugated bilirubin (UCB), and not the total UCB level, has been shown to correlate with bilirubin cytotoxicity, but the key molecular mechanisms accounting for the toxic effects of UCB are largely unknown. Findings Mouse liver mitochondria increase unbound UCB oxidation, consequently increasing the apparent rate constant for unbound UCB oxidation by HRP (Kp), higher than in control and mouse brain mitochondria, emphasizing the importance of determining Kp in complete systems containing the organelles being studied. The in vitro effects of UCB on cytochrome c oxidase activity in mitochondria isolated from mouse brain and liver were studied at Bf ranging from 22 to 150 nM. The results show that UCB at Bf up to 60 nM did not alter mitochondrial cytochrome c oxidase activity, while the higher concentrations significantly inhibited the enzyme activity by 20% in both liver and brain mitochondria. Conclusions We conclude that it is essential to include the organelles being studied in the medium used in measuring both Kp and Bf. A moderately elevated, pathophysiologically-relevant Bf impaired the cytochrome c oxidase activity modestly in mitochondria from mouse brain and liver.