Renal defects in KCNE1 knockout mice are mimicked by chromanol 293B in vivo: identification of a KCNE1-regulated K+ conductance in the proximal tubule.

KCNE1 is a protein of low molecular mass that is known to regulate the chromanol 293B and clofilium-sensitive K+ channel, KCNQ1, in a number of tissues. Previous work on the kidney of KCNE1 and KCNQ1 knockout mice has revealed that these animals have different renal phenotypes, suggesting that KCNE1 may not regulate KCNQ1 in the renal system. In the current study, in vivo clearance approaches and whole cell voltage-clamp recordings from isolated renal proximal tubules were used to examine the physiological role of KCNE1. Data from wild-type mice were compared to those from KCNE1 knockout mice. In clearance studies the KCNE1 knockout mice had an increased fractional excretion of Na+, Cl−, HCO3(−) and water. This profile was mimicked in wild-type mice by infusion of chromanol 293B, while chromanol was without effect in KCNE1 knockout animals. Clofilium also increased the fractional excretion of Na+, Cl− and water, but this was observed in both wild-type and knockout mice, suggesting that KCNE1 was regulating a chromanol-sensitive but clofilium-insensitive pathway. In whole cell voltage clamp recordings from proximal tubules, a chromanol-sensitive, K+-selective conductance was identified that was absent in tubules from knockout animals. The properties of this conductance were not consistent with its being mediated by KCNQ1, suggesting that KCNE1 regulates another K+ channel in the renal proximal tubule. Taken together these data suggest that KCNE1 regulates a K+-selective conductance in the renal proximal tubule that plays a relatively minor role in driving the transport of Na+, Cl− and HCO3(−).

A novel dephosphorylation-activated conductance in a mouse renal collecting duct cell line.

Autosomal dominant polycystic kidney disease (ADPKD) is one of the most common inherited renal diseases. It is associated with the progressive development of renal tubular cysts, which may subsequently lead to renal failure. Studies into the genetic basis of ADPKD have identified two genes, PKD1 and PKD2, that are mutated in ADPKD patients. The PKD1 and PKD2 genes encode for two different proteins, TRPP1 and TRPP2. Previous studies have demonstrated the presence of both TRPP1 and TRPP2 in the renal collecting duct cell line M8. The aim of the following study was to investigate the functional properties of cation currents in these cells and to examine the effect of overexpression of TRPP1 using a transgenic cell model (M7). In M8 cells, initial whole cell currents were low. However, over time there was activation of a flow-sensitive current, which was inhibited by gadolinium (I(Gd)). The I(Gd) was more selective for cations over anions, but did not discriminate between monovalent cations and was Ca2+ permeable. Activation of I(Gd) was dependent on the presence of Ca2+ and also required dephosphorylation. The protein phosphatase 2A inhibitor okadaic acid prevented activation of I(Gd), suggesting that protein phosphatase 2A plays an important role in channel activation. The properties and magnitude of I(Gd) were unaffected in M7 cells, suggesting that overexpression of TRPP1 was without effect. I(Gd) was selectively inhibited by an antibody raised against the C-terminus of TRPP2. However, its selectivity profile was different to TRPP2, suggesting that it is attributable to a TRPP2-like channel or a TRPP2-containing heteromeric channel. In conclusion, these data describe the functional identification of a novel dephosphorylation- and flow-activated TRPP2-related channel in mouse collecting duct cells.

A Kir2.3-like K+ conductance in mouse cortical collecting duct principal cells.

K(+) channels play an important role in renal collecting duct cell function. The current study examined barium (Ba(2+))-sensitive whole-cell K(+) currents (IKBa) in mouse isolated collecting duct principal cells. IKBa demonstrated strong inward rectification and was inhibited by Ba(2+) in a dose- and voltage-dependent fashion, with the K (d) decreasing with hyperpolarization. The electrical distance of block by Ba(2+) was around 8.5%. As expected for voltage-dependent inhibition, the association constant increased with hyperpolarization, suggesting that the rate of Ba(2+) entry was increased at negative potentials. The dissociation constant also increased with hyperpolarization, consistent with the movement of Ba(2+) ions into the intracellular compartment at negative potentials. These properties are not consistent with ROMK but are consistent with the properties of Kir2.3. Kir2.3 is thought to be the dominant basolateral K(+) channel in principal cells. This study provides functional evidence for the expression of Kir2.3 in mouse cortical collecting ducts and confirms the expression of Kir2.3 in this segment of the renal tubule using reverse-transcriptase polymerase chain reaction. The conductance described here is the first report of a macroscopic K(+) conductance in mouse principal cells that shares the biophysical profile of Kir2.3. The properties and dominant nature of the conductance suggest that it plays an important role in K(+) handling in the principal cells of the cortical collecting duct.

Adaptive downregulation of a quinidine-sensitive cation conductance in renal principal cells of TWIK-1 knockout mice.

TWIK-1, a member of the two-pore domain K(+) channel family, is expressed in brain, kidney, and lung. The aim of this study was to examine the effect of loss of TWIK-1 on the renal cortical collecting duct. Ducts were isolated from wild-type and TWIK-1 knockout mice by enzyme digestion and whole-cell clamp obtained via the basolateral membrane. Current- and voltage-clamp approaches were used to examine K(+) conductances. No difference was observed between intercalated cells from wild-type or knockout ducts. In contrast, knockout principal cells were hyperpolarized compared to wild-type cells and had a reduced membrane conductance. This was a consequence of a fall in a barium-insensitive, quinidine-sensitive conductance (G (Quin)). G (Quin) demonstrated outward rectification and had a relatively low K(+) to Na(+) selectivity ratio. Loss of G (Quin) would be expected to lead to the hyperpolarization observed in knockout ducts by increasing fractional K(+) conductance and Na(+) uptake by the cell. Consistent with this hypothesis, knockout ducts had an increased diameter in comparison to wild-type ducts. These data suggest that G (Quin) contributes to the resting membrane potential in the cortical collecting duct and that a fall in G (Quin) could be an adaptive response in TWIK-1 knockout ducts.

Euglycemic clamp study in clozapine-induced diabetic ketoacidosis.

OBJECTIVE: To describe the fifth case of clozapine-induced diabetic ketoacidosis (DKA) with complete resolution of abnormal glucose metabolism after discontinuation of clozapine as assessed by oral glucose tolerance testing (OGTT) and the first to be serially studied with markers of pancreatic autoimmunity; to demonstrate insulin resistance using the euglycemic clamp study and reduced pancreatic insulin reserve using intravenous glucose tolerance testing (IVGTT) in clozapine-induced diabetes mellitus and DKA, when the OGTT was normal; and to systematically review the previously described cases of clozapine-induced diabetes mellitus and DKA. CASE SUMMARY: A 33-year-old white man without past or family history of diabetes mellitus presented with DKA after eight months of clozapine therapy (50 mg twice daily). After treatment of DKA and discontinuation of clozapine, glucose tolerance and concurrent serum insulin concentrations reverted to normal as measured by two OGTT performed 60 and 320 days after resolution of DKA. DISCUSSION: Antiislet-cell antibodies, antiglutamic acid decarboxylase antibodies, and human insulin antibody were negative on two separate occasions. Euglycemic clamp study demonstrated insulin resistance manifested by a glucose disposal rate of approximately 55% of mean normal values. IVGTT demonstrated a low rate of glucose disappearance (KG = 0.95) and diminished first-phase insulin response when OGTT was normal, indicating impairment in insulin sensitivity and reduction in beta cell function 323 days after discontinuance of clozapine. This adverse reaction is considered probable according to the Naranjo probability scale. CONCLUSIONS: The occurrence of cases of DKA and new or worsening diabetes mellitus in patients using clozapine suggests a causal relationship. We hypothesize that the mechanism by which clozapine may produce glucose intolerance may require a preexisting latent defect in insulin secretion and insulin action. With the administration of clozapine, some of these patients may develop worsening insulin resistance and may fail to mount an appropriate compensatory beta cell insulin secretion for the degree of insulin resistance. As a consequence, hyperglycemia develops and its persistence results in glucose toxicity, further suppressing beta cell insulin secretion. Such combined defects in insulin secretion and sensitivity are known to be synergistic, leading to the development of abnormal glucose tolerance, which can be clinically manifested as a spectrum ranging from impaired glucose tolerance through severe hyperglycemia to DKA. Patients being started on clozapine should be carefully followed for the development or worsening of diabetes mellitus, regardless of the dose of the drug.

Type 1 aldosterone synthase deficiency presenting in a middle-aged man.

Aldosterone synthase deficiency due to mutations in the CYP11B2 gene usually presents in infancy with electrolyte abnormalities and failure to thrive, whereas affected adults are usually asymptomatic. We describe a patient who first came to medical attention in middle age when he developed hyperkalemia after preparation for a barium enema. Past medical history was notable for failure to thrive in infancy. He had elevated PRA with low serum and urinary levels of aldosterone and its metabolites and normal or slightly elevated levels of 18-hydroxycorticosterone. These findings suggested a diagnosis of type 1 aldosterone synthase deficiency. The patient had a homozygous duplication of six nucleotides at codon 143 in exon 3 of CYP11B2, leading to the insertion of two amino acid residues (Arg-Leu). When the corresponding mutant complementary DNA was expressed in cultured cells, the resulting enzyme was completely inactive, confirming the diagnosis. We conclude that aldosterone synthase deficiency represents an unusual cause of hyperreninemic hypoaldosteronism presenting in adult life, but it should be suspected if the past medical history is positive for failure to thrive in childhood or if the patient manifests no other recognized causes of hyperreninemic hypoaldosteronism.

Diffuse stromal Leydig cell hyperplasia: a unique cause of postmenopausal hyperandrogenism and virilization.

A 60-year-old woman presented with diffuse scalp alopecia, hirsutism, and clitorimegaly, and the mean serum testosterone levels were greater than 200 ng/dL. Findings on computed tomography of both adrenal glands were normal. After bilateral oophorectomy, a unique histological picture consisting of diffuse stromal Leydig cell hyperplasia was found. Reinke crystals were present, but neither hilus cell hyperplasia nor stromal hyperthecosis was noted. Sequencing of the 11 exons of the gene for the luteinizing hormone receptor revealed no abnormality. Relevant data suggest that treatment of the postmenopausal woman with hyperandrogenism and virilization is bilateral laparoscopic oophorectomy if she has no pronounced ovarian enlargement or adrenal tumor on imaging. In this setting, an intensive endocrine evaluation or a search for metastatic disease seems to be unnecessary.

Syndrome of inappropriate antidiuretic hormone secretion associated with lisinopril.

OBJECTIVE: To describe a case of the syndrome of inappropriate antidiuretic hormone secretion (SIADH) associated with lisinopril therapy. CASE SUMMARY: A 76-year-old white woman who was being treated with lisinopril and metoprolol for hypertension presented with headaches accompanied by nausea and a tingling sensation in her arms. Her serum sodium was 109 mEq/L, with a serum osmolality of 225 mOsm/kg, urine osmolality of 414 mOsm/kg, and spot urine sodium of 122 mEq/L. Diclofenac 75 mg qd for osteoarthritic pain and lisinopril 10 mg qd for hypertension was begun in 1990. Lisinopril was increased to 20 mg qd in August 1994 and to 20 mg bid pm in August 1996 for increasing blood pressure; metoprolol 50 mg qd was added in July 1996. A diagnosis of SIADH was postulated and further evaluation was undertaken to exclude thyroid and adrenal causes. After lisinopril was discontinued and the patient restricted to 1000 mL/d of fluid, serum sodium gradually corrected to 143 mEq/L. The patient was discharged taking metoprolol alone for her hypertension; serum sodium has remained > or =138 mEq/L through April 1999, 32 months after discharge, despite daily use of diclofenac. DISCUSSION: Angiotensin-converting enzyme (ACE) inhibitors in antihypertensive doses may block conversion of angiotensin I to angiotensin II in the peripheral circulation, but not in the brain. Increased circulating angiotensin I enters the brain and is converted to angiotensin II, which may stimulate thirst and release of antidiuretic hormone from the hypothalamus, eventually leading to hyponatremia. CONCLUSIONS: SIADH should be considered a rare, but possible, complication of therapy with lisinopril and other ACE inhibitors.

Nephrogenic diabetes insipidus persisting 57 months after cessation of lithium carbonate therapy: report of a case and review of the literature.

OBJECTIVE: To illuminate the natural history of prolonged nephrogenic diabetes insipidus after discontinuation of lithium carbonate treatment and to assess the response to therapy with desmopressin acetate and triamterene-hydrochlorothiazide. METHODS: We analyzed sequential determinations of serum and urine osmolality, plasma arginine vasopressin, serum sodium, blood urea nitrogen, calcium, ionized calcium, parathyroid hormone, and 24-hour urine volume during a period of 57 months in a 67-year-old woman. RESULTS: Our patient experienced persistent polyuria in conjunction with having repeated serum osmolalities between 300 and 323 mOsm/kg and urine osmolalities between 130 and 208 mOsm/kg. Concomitant plasma arginine vasopressin levels were as high as 12.0 pg/mL, consistent with the diagnosis of nephrogenic diabetes insipidus. Administration of triamterene-hydrochlorothiazide reduced 24-hour urine volume and serum osmolality while increasing urine osmolality. Desmopressin acetate exhibited no effect. CONCLUSION: In this report, we describe the eighth documented case of persistent nephrogenic diabetes insipidus, lasting 57 months after cessation of lithium therapy, and demonstrate a palliative effect of triamterene-hydrochlorothiazide.

Assessment of inner dynein arm structure and possible function in ciliary and flagellar axonemes.

The construction and assessment of a three-dimensional computer-generated model of inner dynein arms on a 96-nm repeat unit of an axonemal doublet is described. The model is based on published electron micrographs of axonemes from Tetrahymena cilia and eel sperm, which were prepared using several different techniques: negative stain, freeze etch, and thin section. The inner arm structure is represented as three inner dynein arm complexes containing four inner dynein arms (IDAs), three dyads, and one single-headed arm, each capable of bridging the interdoublet gap. The IDA structures in the model have been correlated with the domains containing dynein heavy-chain isoforms mapped by several authors using genetic analyses of Chlamydomonas mutants. The model is consistent with micrographic evidence from axonemes of cilia and flagella from other organisms that led previously to conflicting structural interpretations. In this reconciling interpretation, the different alignments of the IDAs relative to the corresponding outer dynein arms observed in micrographs of differently prepared samples, result from the IDAs being arrested at different stages of their cycles of activity in each preparation. By interpolating between these positions of arrest, cycles of activity are proposed for each of the IDAs during which the arms attach to the neighbouring doublet microtubule and drive it tipwards.

Histologically proven lymphocytic hypophysitis: spontaneous resolution and subsequent pregnancy.

Of the 128 previously reported cases of lymphocytic hypophysitis, the diagnosis was histologically proven in 6 patients in whom the pituitary mass regressed spontaneously; only 1 subsequently became pregnant. Among six additional patients who became pregnant after a diagnosis of presumed lymphocytic hypophysitis, the disease was confirmed histologically in only three, two of whom underwent surgical debulking and one who had no follow-up imaging. To our knowledge, we describe the second patient with histologically proven lymphocytic hypophysitis, associated with adrenocorticotropic hormone (corticotropin) and prolactin deficiencies, in whom the pituitary mass regressed completely with physiologic hydrocortisone therapy only and in whom a spontaneous pregnancy occurred subsequently with no postpartum recurrence of the pituitary mass. This information lends credence to previous recommendations that, in the absence of visual field defects, surgical and corticosteroid therapy may be safely withheld, with periodic reassessment.

Visualizing and quantifying molecular goodness-of-fit: small-probe contact dots with explicit hydrogen atoms.

The technique of small-probe contact dot surfaces is described as a method for calculating and displaying the detailed atomic contacts inside or between molecules. It allows one both to measure and to visualize directly the goodness-of-fit of packing interactions. It requires both highly accurate structures and also the explicit inclusion of all hydrogen atoms and their van der Waals interactions. A reference dataset of 100 protein structures was chosen on the basis of resolution (1.7 A or better), crystallographic R-value, non-homology, and the absence of any unusual problems. Hydrogen atoms were added in standard geometry and, where needed, with rotational optimization of OH, SH, and NH+3 positions. Side-chain amide orientations were corrected where required by NH van der Waals clashes, as described in the accompanying paper. It was determined that, in general, methyl groups pack well in the default staggered conformation, except for the terminal methyl groups of methionine residues, which required rotational optimization. The distribution of serious clashes (i.e. non-H-bond overlap of >/=0.4 A) was studied as a function of resolution, alternate conformations, and temperature factor (B), leading to the decision that packing and other structural features would not be analyzed for residues in 'b' alternate conformations or with B-factors of 40 or above. At the level of the fine details analyzed here, structural accuracy improves quite significantly over the range from 1.7 to 1.0 A resolution. These high-resolution structures show impressively well-fitted packing interactions, with some regions thoroughly interdigitated and other regions somewhat sparser. Lower-resolution structures or model structures could undoubtedly be improved in accuracy by the incorporation of this additional information: for example, nucleic acid structures in non-canonical conformations are often very accurate for the bases and much less reliable for the backbone, whose conformation could be specified better by including explicit H atom geometry and contacts. The contact dots are an extremely sensitive method of finding problem areas, and often they can suggest how to make improvements. They can also provide explanations for structural features that have been described only as empirical regularities, which is illustrated by showing that the commonest rotamer of methionine (a left-handed spiral, with all chi values near -60 degrees) is preferred because it provides up to five good H atom van der Waals contacts. This methodology is thus applicable in two different ways: (1) for finding and correcting errors in structure models (either experimental or theoretical); and (2) for analyzing interaction patterns in the molecules themselves.

Hypercalcemia associated with infection by Cryptococcus neoformans and Coccidioides immitis.

BACKGROUND: Of the 13 reported cases of hypercalcemia associated with fungal infection, 1 was caused by Cryptococcus neoformans and probably mediated by increased levels of 1,25-dihydroxyvitamin D [1,25(OH)2D]. Eight others were associated with Coccidioides immitis, of which only 2 had measured 1,25(OH)2D levels; in both, they were diminished. We report a patient with human immunodeficiency virus infection and simultaneous C. immitis and C. neoformans pneumonia and C. immitis fungemia associated with hypercalcemia. METHODS: Consecutive measurements of serum total and ionized calcium, phosphorous, blood urea nitrogen, creatinine, 25(OH)D, 1,25(OH)2D, parathyroid hormone (PTH), parathyroid hormone-related protein (PTHrp) and albumin were performed over a period of 46 months. RESULTS: While the patient was hypercalcemic, intact serum PTH and PTHrp were undetectable, serum 25(OH)D levels were normal, and serum 1,25(OH)2D levels were in the high normal range. Successful treatment of the C. immitis and C. neoformans infections resulted in resolution of the hypercalcemia and increase of PTH and PTHrp to the normal range. CONCLUSION: In some patients with HIV infection, coincident hypercalcemia, and severe fungal infection, the responsible factor may be 1,25(OH)2D. Although total serum levels of this compound may not be frankly elevated, they are inappropriately high for the circumstances. Free 1,25(OH)2D levels should be determined in this situation.

Transient ionized hypocalcemia and secondary hyperparathyroidism accompanying acute adrenal insufficiency.

OBJECTIVE: To describe the hitherto unrecognized occurrence of transient ionized hypocalcemia with acute adrenal insufficiency and its therapy. METHODS: We present three case reports with documented longitudinal laboratory findings. RESULTS: Transient ionized hypocalcemia of acute illness has been noted in children and adults and is associated with increased mortality. Precipitating illnesses include gram-positive and gram-negative sepsis and staphylococcal toxic shock syndrome. We encountered three patients with transient ionized hypocalcemia associated with acute adrenal insufficiency. Similar to severely ill, transiently hypocalcemic patients without adrenal insufficiency, one patient demonstrated 25-hydroxyvitamin D deficiency, a second had minimal magnesium deficiency, and a third had no identifiable underlying abnormality. All three patients exhibited a transient increase in levels of serum intact parathyroid hormone and 1,25-dihydroxyvitamin D in response to ionized hypocalcemia, indicative of temporary secondary hyperparathyroidism. Two of the three patients were treated solely with glucocorticoids and intravenous administration of fluids, whereas the third received minimal intramuscularly administered magnesium and antibiotics in addition. All ultimately demonstrated a return to normal of serum total and ionized calcium, parathyroid hormone, and 1,25-dihydroxyvitamin D with no further treatment, even though one patient remained deficient in 25-hydroxyvitamin D. CONCLUSION: On the basis of these cases, we conclude that acute adrenal insufficiency and its treatment must be added to the disorders associated with transient ionized hypocalcemia and that transient secondary hyperparathyroidism is characteristic of at least some of the patients.

Design and calibration of electric field probes in the range 10-120 MHz.

In view of potential thermal hazards, there is a need to determine the specific absorption rate (SAR) distributions associated with radiofrequency coils used in magnetic resonance imaging (MRI) (typically 10-120 MHz). Electric field (E-field) distributions in tissue-equivalent phantoms may be determined using a probe comprising a dipole antenna and a detector. The geometry of the dipole dictates the sensitivity of the device, thus two designs are discussed in this paper. Both probes are compact, have a spatial resolution of 2.5 cm3, operate at MR frequencies and have a response independent of the dielectric characteristics of the phantom material. Calibration of these probes requires a system capable of producing a known E-field both in air and in a tissue-like medium at frequencies between 10 and 120 MHz. Transverse electromagnetic wave (TEM) cells answering these specifications are described and the calibration procedure outlined. Accurately calibrated E-field probes can make field measurements in phantoms which can be used to verify predictions from numerical models. These numerical techniques may then be used to predict E-fields, and hence SAR, in patients.

Experimental verification of numerically predicted electric field distributions produced by a radiofrequency coil.

There are safety issues regarding energy deposition within tissues due to radiofrequency fields used in some magnetic resonance (MR) procedures. Procedures should be compliant with guidelines that specify limits to temperature elevation and specific absorption rate (SAR). In general, direct measurement of these quantities in patients is impractical and an alternative approach is to determine SAR from the electric field (E-field) distributions predicted by numerical models. In this initial study the E-field distribution in a tissue-simulating phantom due to a square coil driven at 31 MHz is predicted using a finite-difference time domain (FDTD) solution to Maxwell's equations. An experimental arrangement of the same problem was constructed and the resulting E-field distribution was measured using a calibrated minimally perturbing E-field probe. A comparison between experimentally and theoretically derived data showed that the numerically predicted E-fields were within +/-1 dB of the fields measured with the E-field probe in the phantom material. The results provide confidence in the use of the FDTD algorithm to determine quantitatively accurate E-field distributions arising from square radiofrequency (RF) coils used in MR procedures. The accuracy of numerical models of other coil designs such as bird cages, saddles and surface coils can be investigated in the same manner. Future studies will evaluate the exposure of patients to these RF fields.

Congenital adrenal hyperplasia presenting as massive adrenal incidentalomas in the sixth decade of life: report of two patients with 21-hydroxylase deficiency.

Divergent recommendations exist regarding the evaluation of adrenal incidentalomas. Recent data have indicated a prevalence of adrenal tumors of 71% in nonclassical congenital adrenal hyperplasia (CAH) and unmasked heterozygotes. These data expand the differential diagnosis of such incidental tumors and substantially modify the approach to their evaluation. We present two patients, female pseudohermaphrodites with the simple virilizing form of CAH and 21-hydroxylase deficiency, who functioned successfully as married phenotypic males. Both came to medical attention in the sixth decade by virtue of massive adrenal incidentalomas encountered in the evaluation of recurrent urinary tract infections. Each had a 46, XX karyotype, no palpable testes, and markedly elevated baseline levels of 17-hydroxyprogesterone (17-OH Prog) of 6086 ng/dL and 6750 ng/dL. Both responded appropriately to dexamethasone suppression with reduction of 17-OH Prog, androgens and, in the second patient, ACTH to normal or near normal levels. Histologic and autopsy examination of the first patient's tumor and computed tomographic characteristics of the second revealed a benign adenoma and myelolipoma respectively. We extend and confirm previous recommendations that CAH be included in the differential diagnosis of adrenal incidentaloma and that baseline 17-OH Prog. levels be obtained, with ACTH stimulation if necessary, to diagnose the presence of nonclassical CAH.

Distinct regulatory control of the Brachyury gene in axial and non-axial mesoderm suggests separation of mesoderm lineages early in mouse gastrulation.

Brachyury is required for the normal extension of the anteroposterior axis during mouse embryogenesis. A transgene comprising sequences from -500 to +150 relative to the start of Brachyury transcription, and the reporter gene lacZ, recapitulates some, but not all elements of Brachyury expression. Beta-Galactosidase expression is seen in the primitive streak from 6.5 d.p.c. but there is no detectable reporter expression in the node or notochord. Thus, the regulatory sequences required for the expression of Brachyury in the cells traversing the primitive streak are distinct from those required for the initiation of expression in the node. This suggests that different or additional signals are involved in activation of Brachyury in the node and notochord than those inducing Brachyury in the primitive streak. Additionally, the data suggest the possibility that axial and non-axial mesoderm are distinct from the earliest stages of Brachyury expression.

Intrahepatic cholestasis during nicotinic acid therapy.

BACKGROUND: Nicotinic acid, widely used to lower serum cholesterol levels, may rarely cause cholestatic jaundice. SUMMARY: A 61-year-old white man with hypercholesterolemia complained of marked pruritus and became jaundiced after taking 3.0 g of crystalline nicotinic acid daily for 13 months. His total serum bilirubin level was increased at 144 mumol/L (8.4 mg/dL) and his alkaline phosphatase level was markedly elevated at 35.00 mukat/L (2100 U/L). Endoscopic retrograde cholangiopancreatography failed to demonstrate an obstructive lesion in the extrahepatic biliary system, computed tomography showed no intrahepatic dilatation, and ultrasonographic studies of the liver, gallbladder, and pancreas were normal; these factors all suggest intrahepatic cholestasis. Symptoms improved and liver function test results returned to normal within 51 days after stopping the drug. CONCLUSIONS: Nicotinic acid-induced cholestatic jaundice may not be as rare as previously thought, and physicians should observe their patients for it.

Control of the effective P/O ratio of oxidative phosphorylation in liver mitochondria and hepatocytes.

The control exerted by substrate oxidation reactions, by ATP turnover and by the proton leak over the oxygen consumption rate, the phosphorylation rate, the proton leak rate and the protonmotive force (delta p) in isolated rat liver mitochondria under a range of conditions between non-phosphorylating (State 4) and maximum phosphorylation (State 3) was investigated by using the top-down approach of metabolic control analysis. The experiments were carried out with saturating concentrations of the substrates succinate, glutamate with malate, or pyruvate with malate. The distribution of control was very similar with each of the three substrates. The effective P/O ratio (i.e. not corrected for leak reactions) was also measured; it varied from zero in State 4 to 80-90% of the maximum theoretical P/O ratio in State 3. Under most conditions control over the effective P/O ratio was shared between proton leak (which had negative control) and the phosphorylating subsystem (which had roughly equal positive control); near State 4, substrate oxidation reactions also acquired some control over this ratio. In resting hepatocytes the effective P/O ratio was only 50% of its maximum theoretical value, corresponding to an effective P/O ratio of only 1.3 for complete oxidation of glucose. The effective P/O ratio for intracellular mitochondrial oxygen consumption was 64% of the maximum value. The control coefficient of the mitochondrial proton leak over the effective P/O ratio in cells was -0.34; the control coefficient of phosphorylation reactions over this ratio was 0.31 and the control coefficient of substrate oxidation reactions over the ratio was 0.03, showing how the coupling efficiency in cells can respond sensitively to agents that change the proton leak or the ATP demand, but not to those that change substrate oxidation.