LOW PREVALENCE OF HAEMOSPORIDIANS IN BLOOD AND TISSUE SAMPLES FROM HUMMINGBIRDS.

Hummingbirds are vital members of terrestrial ecosystems, and because of their high metabolic requirements, they serve as indicators of ecosystem health. Monitoring the parasitic infections of hummingbirds is thus especially important. Haemosporidians, a widespread group of avian blood parasites, are known to infect hummingbirds, but little is known about the prevalence and diversity of these parasites in hummingbirds. The prevalence of haemosporidians in several hummingbird species was examined and we compared 4 different tissue types in detecting parasites by polymerase chain reaction (PCR). Blood samples from 339 individuals of 3 different hummingbird species were tested, and 4 individuals were found positive for haemosporidian infection, a prevalence of 1.2%. Hummingbird carcasses (n = 70) from 5 different hummingbird species were also sampled to assess differences in detection success of haemosporidians in heart, kidney, liver, and pectoral muscle tissue samples. Detection success was similar among tissue types, with haemosporidian prevalence of 9.96% in heart tissue, 9.52% in kidney tissue, 10.76% in liver tissue, and 11.76% in pectoral muscle tissue. All tissue samples positive for haemosporidian infection were from the Black-chinned Hummingbird (Archilochus alexandri). Possible reasons for low prevalence of these blood parasites could include low susceptibility to insect vectors or parasite incompatibility in these hummingbirds.

Dynamic measurements of the acute and chronic effects of lysosomotropic agents on hepatocyte lysosomal pH using flow cytometry.

Previous investigators measuring the pH of lysosomes have used digitized video microscopy (DVM) in freshly isolated or cultured cells. Although useful, this technique is time consuming, requires the use of an image analysis system, and is limited by the fact that measurements can be made in only a relatively small number of cells. The aim of this study was to develop and initially apply a technique using flow cytometry to make dynamic measurements of lysosomal pH in a large number of living hepatocytes. Rats were injected intraperitoneally with fluorescein isothiocyanate-dextran (FITC-Dex), a pH-sensitive fluorescent probe that is sequestered into lysosomes. Hepatocytes were isolated 16 hours after injection by collagenase perfusion. Lysosomal pH was measured in 20,000 hepatocytes per animal using flow cytometry with excitation at 488 nm and emission at 530 nm (pH sensitive) and 585 nm (pH insensitive). A standard curve of pH versus the 530/585 nm ratio was generated with FITC-Dex-loaded hepatocytes by equilibrating intralysosomal pH with extracellular pH using ionophores and metabolic inhibitors. The acute effects of chloroquine and methylamine were determined by exposing isolated hepatocytes to these lysosomotropic agents. The effect of chronic administration of chloroquine and Triton WR-1339 (Rutger Chemical, Inc., Irvington, NJ) on lysosomal pH was also measured. Intralysosomal pH was 4.67 + 0.02, nearly identical to the value 4.70 + 0.05 previously measured by us using DVM. Both chloroquine and methylamine caused both rapid (< 1 minute), major (0.5 to 2.0 pH units), and dose-dependent increases in lysosomal pH as well as changes in lysosome morphology.(ABSTRACT TRUNCATED AT 250 WORDS)

Plasma glutathione concentration in patients with chronic hepatitis C virus infection.

It has recently been proposed that a depletion of glutathione (GSH) may be a contributing factor to viral persistence and resistance to interferon-alpha (IFN-alpha) therapy in chronic hepatitis C virus (HC) infection. The aim of this study was: (1) to compare plasma GSH levels in patients with chronic HCV infection and normal healthy controls; and (2) to correlate GSH levels with liver histology and serum HCV RNA levels. Twenty-four patients with compensated chronic hepatitis C and 27 healthy subjects were studied. Serum and heparinized plasma were prospectively prepared and frozen within 1 h of collection. Plasma glutathione and glutathione peroxidase (GP) levels were measured spectrophotometrically. The serum HCV RNA level was quantitated by the branched chain DNA signal-amplification assay. Plasma GSH levels were not decreased in patients with chronic HCV infection but were actually greater than in controls (control 1.27 +/- 0.12 micrograms ml-1, HCV 1.62 +/- 0.11 micrograms ml-1, P < 0.05). There was also no difference in plasma GP activity between these two groups (control 0.233 +/- 0.007 U ml-1, HCV 0.230 +/- 0.007 U ml-1). Among the patients with chronic HCV infection, there was no correlation between either plasma GSH or GP levels and the serum alanine aminotransferase (ALT) or aspartate aminotransferase (AST), serum HCV RNA level, or liver histology. This study demonstrates that chronic HCV infection does not decrease the plasma GSH and GP levels.

Hepatic infarction complicating a transjugular intrahepatic portosystemic shunt.

Transjugular intrahepatic portocaval shunt (TIPS) is being used with increasing frequency to treat variceal bleeding and portal hypertension. Like any other invasive vascular procedure, it has the potential for serious complications. We report a case of a woman who underwent a TIPS procedure for variceal bleeding due to hepatitis C and sustained a hepatic infarction as a complication of the procedure. We caution about the potential for catastrophic complications with this procedure, and recommend that it be performed only in centers with a liver transplant program nearby.

Alterations in hepatocyte lysosomes in experimental hepatic copper overload in rats.

BACKGROUND: Although Wilson's disease is characterized by an accumulation of copper within hepatocyte lysosomes, the effects of excess copper on hepatic lysosomes are unknown. We studied the effects of excess copper on the structure, physicochemical properties, and pH of hepatocyte lysosomes using a rodent model. METHODS: Rats were copper loaded with 0.125% copper acetate in water for 6 weeks. Copper was measured by atomic absorption spectrophotometry. Morphology was studied by electron microscopy. Lysosomal membrane fluidity was studied by fluorescence polarization, and lipid composition was determined by gas chromatography. Hepatocyte lysosomal pH was determined by flow cytometry. RESULTS: Copper overload resulted in a 10-fold increase in hepatic copper. Hepatocyte lysosomes were enlarged and abnormally shaped with a 27-fold increase in copper, increased in vitro fragility, and decreased lysosomal membrane fluidity. Thiobarbituric acid reactive substances, a measure of lipid peroxidation, doubled in isolated lysosomal membranes. Polyunsaturated fatty acids increased, saturated fatty acids decreased, and membrane content of selected fatty acids was modified after copper overload. Lysosomal pH increased from 4.67 +/- 0.02 to 4.87 +/- 0.02. CONCLUSIONS: Copper overload causes alterations in lysosomal morphology, increases lysosomal fragility, decreases membrane fluidity, alters membrane fatty acid composition, and increases lysosomal pH. Copper catalyzed lipid peroxidation represents the likely mechanism for these alterations.

Relationship of the polymerase chain reaction for cytomegalovirus to the development of hepatitis in liver transplant recipients.

In a pilot study, the polymerase chain reaction was found to be more sensitive than standard viral culture methods for the detection of cytomegalovirus, particularly from blood and tissues. We therefore applied this technique to 71 serially collected liver biopsies from 16 orthotopic liver transplant patients. All patients were CMV-seropositive (n = 15) or seroconverted (n = 1). Seven patients (9 biopsies) had histologically proved CMV hepatitis, and all these biopsies were CMV PCR-positive. Six of these 7 patients had a prior liver biopsy that was CMV PCR-positive, but culture and histology-negative, an average of 13.2 +/- 6.9 days before the histologically positive biopsy. The 7th patient was not biopsied prior to the diagnostic biopsy. Three patients had 7 liver biopsies that were CMV PCR-positive, but histologically negative for CMV hepatitis. Two of these three had CMV infection confirmed by viral culture of blood or liver biopsy. The remaining 6 patients had a total of 26 liver biopsies that were negative for CMV by PCR, culture, and histology. Among liver transplant patients, CMV PCR performed on liver biopsy specimens correctly identified all histologically proven cases of CMV hepatitis. CMV PCR positivity in liver tissue did not correlate with latent infection and preceded the development of CMV hepatitis or other meaningful CMV infection in 8 of 10 patients.

Treatment of colonic bleeding in Klippel-Trenaunay syndrome with combined partial colectomy and endoscopic laser.

Klippel-Trenaunay syndrome can be associated with significant lower gastrointestinal bleeding from diffuse cavernous hemangiomata of the colon. Distal colectomy is curative but the formation of a colostomy is undesirable in patients who already have a poor self-image due to the other manifestations of Klippel-Trenaunay syndrome such as multiple cutaneous hemangiomata, severe varicose veins, and prominent limb hypertrophy. Endoscopic laser ablation alone is often not feasible in these patients because of the extensive and often circumferential nature of the colonic hemangiomata. This report describes successful treatment of gastrointestinal hemorrhage due to colonic hemangiomata by a novel combination of partial colonic resection and endoscopic laser therapy. Case 1 demonstrates that endoscopic laser ablation of the diseased colonic mucosa, while not feasible with the extensive involvement of the left colon, was very effective when combined with partial colectomy at curing the bleeding while maintaining anal sphincter function. In case 2 the disease was more limited and more easily amenable to endoscopic laser ablation. Combined surgical and laser endoscopic therapy is recommended in treating the colonic hemangiomata associated with Klippel-Trenaunay syndrome.

Gastrointestinal involvement in a woman with dyskeratosis congenita.

Dyskeratosis congenita (DC) is a rare hereditary condition with characteristic dermatological findings and is frequently associated with the development of pancytopenia. Involvement of the gastrointestinal tract and liver occurs in a significant proportion of patients. Although the disorder is transmitted in an X-linked fashion, several affected females have been described. We report a case of DC involving a woman in whom gastrointestinal symptoms were prominent, primarily diarrhea, gastrointestinal hemorrhage, and ascites. Portal hypertension was evident by the presence of splenorenal varices. Extensive evaluation failed to disclose an etiology for either her colitis or portal hypertension. The literature concerning gastrointestinal involvement in DC is reviewed and a possible association with noncirrhotic portal hypertension is suggested.

Cytomegalovirus as a cause of isolated severe ileal bleeding.

A 66-year-old man with squamous cell carcinoma of the tongue and vallecula had massive lower gastrointestinal hemorrhage. Colonoscopy examination results were normal, but angiography revealed a 3 x 6-cm hypervascular mass in the right lower quadrant. Persistent bleeding required resection of a large ileal ulcer, which proved to be an isolated ulcer containing cells with cytomegalovirus (CMV) inclusion bodies. Massive gastrointestinal hemorrhage due to an isolated CMV ileal ulcer in adults without acquired immunodeficiency syndrome has not been previously reported. This case illustrates that hemorrhage due to CMV infection of the ileum may occur in the absence of endoscopic evidence of colonic involvement.

Alterations in the structure, physicochemical properties, and pH of hepatocyte lysosomes in experimental iron overload.

While hemochromatosis is characterized by sequestration of iron-protein complexes in hepatocyte lysosomes, little is known about the effects of excess iron on these organelles. Therefore, we studied the effects of experimental iron overload on hepatocyte lysosomal structure, physicochemical properties, and function in rats fed carbonyl iron. A sixfold increase (P less than 0.0001) in hepatic iron and a fivefold increase in lysosomal iron (P less than 0.01) was observed after iron loading; as a result, hepatocyte lysosomes became enlarged and misshapen. These lysosomes displayed increased (P less than 0.0001) fragility; moreover, the fluidity of lysosomal membranes isolated from livers of iron-loaded rats was decreased (P less than 0.0003) as measured by fluorescence polarization. Malondialdehyde, an end product of lipid peroxidation, was increased by 73% (P less than 0.008) in lysosomal membranes isolated from livers of iron-overloaded rats. While amounts of several individual fatty acids in isolated lysosomal membranes were altered after iron overload, cholesterol/phospholipid ratios, lipid/protein ratios, double-bond index, and total saturated and unsaturated fatty acids remained unchanged. The pH of lysosomes in hepatocytes isolated from livers of iron-loaded rats and measured by digitized video microscopy was increased (control, 4.70 +/- 0.05; iron overload, 5.21 +/- 0.10; P less than 0.01). Our results demonstrate that experimental iron overload causes marked alterations in hepatocyte lysosomal morphology, an increase in lysosomal membrane fragility, a decrease in lysosomal membrane fluidity, and an increase in intralysosomal pH. Iron-catalyzed lipid peroxidation is likely the mechanism of these structural, physicochemical, and functional disturbances.

POEMS syndrome with idiopathic flushing mimicking carcinoid syndrome.

POEMS syndrome, a rare multisystem disease, is a variant of osteosclerotic myeloma and is characterized by polyneuropathy, organomegaly, endocrinopathy, monoclonal proteins, and skin changes. Presented herein is a case of POEMS syndrome with flushing. The flushing was intermittent, involving the face and upper third of the trunk, and was associated with hypotension and bronchospasm. Final diagnosis was made by biopsy examination of an axillary lymph node, which showed angiofollicular hyperplasia that stained strongly and selectively for lambda light chains. The patient had most of the typical features of POEMS syndrome but was unique in that her most striking finding was carcinoid-like flushing. The flushing improved with steroid therapy, as did some of the other clinical features of her disease. This case suggests that idiopathic flushing can be added to the skin changes observed in POEMS syndrome.

Biliary copper excretion by hepatocyte lysosomes in the rat. Major excretory pathway in experimental copper overload.

We investigated the hypothesis that lysosomes are the main source of biliary copper in conditions of hepatic copper overload. We used a rat model of oral copper loading and studied the relationship between the biliary output of copper and lysosomal hydrolases. Male Sprague-Dawley rats were given tap water with or without 0.125% copper acetate for up to 36 wk. Copper loading produced a 23-fold increase in the hepatic copper concentration and a 30-65% increase in hepatic lysosomal enzyme activity. Acid phosphatase histochemistry showed that copper-loaded livers contained an increased number of hepatocyte lysosomes; increased copper concentration of these organelles was confirmed directly by both x ray microanalysis and tissue fractionation. The copper-loaded rats showed a 16-fold increase in biliary copper output and a 50-300% increase in biliary lysosomal enzyme output. In the basal state, excretory profiles over time were similar for biliary outputs of lysosomal enzymes and copper in the copper-loaded animals but not in controls. After pharmacologic stimulation of lysosomal exocytosis, biliary outputs of copper and lysosomal hydrolases in the copper-loaded animals remained coupled: injection of colchicine or vinblastine produced an acute rise in the biliary output of both lysosomal enzymes and copper to 150-250% of baseline rates. After these same drugs, control animals showed only the expected increase in lysosomal enzyme output without a corresponding increase in copper output. We conclude that the hepatocyte responds to an increased copper load by sequestering excess copper in an increased number of lysosomes that then empty their contents directly into bile. The results provide direct evidence that exocytosis of lysosomal contents into biliary canaliculi is the major mechanism for biliary copper excretion in hepatic copper overload.

Effect of red pepper and black pepper on the stomach.

Spices have long been implicated as a cause of gastric mucosal injury. We assessed the effects of red and black pepper on the gastric mucosa using double-blind intragastric administration of test meals containing red pepper (0.1-1.5 g) or black pepper (1.5 g) to healthy human volunteers; aspirin (655 mg) and distilled water were used as positive and negative controls, respectively. Serial gastric washes were performed after test meal administration and gastric contents were analyzed for DNA, pepsin, blood, sodium, potassium, parietal cell secretion, and nonparietal cell secretion. Both red pepper and black pepper caused significant increases in parietal secretion, pepsin secretion, and potassium loss. Gastric cell exfoliation (as reflected in DNA loss into gastric contents) was increased after red or black pepper administration; the increase after red pepper administration was dose dependent. Mucosal microbleeding was seen after spice administration and one subject had grossly visible gastric bleeding after both red pepper and black pepper administration. There were no significant differences from control between the test meals, in nonparietal volume, fractional recovery of the gastric secretions, or sodium secretion. Finally, no spice was significantly different from aspirin in any parameter studied; indeed, aspirin was comparable to the higher doses of pepper. The long-term result of daily pepper ingestion is unknown. Whether spices are detrimental, beneficial (e.g., inducing an adaptive cytoprotective response), or have no significant long-term effect on the gastric mucosa is unknown and deserves further study.

The pathophysiology of the ocular microenvironment. II. Copper-induced ocular inflammation and hypotony.

The ocular effects of intravitreally injected copper sulfate solutions were studied in New Zealand white rabbits. These injections resulted in uveitis characterized by prolonged ocular hypotony, increased protein concentrations and decreased ascorbic acid concentrations in both the vitreous and aqueous humors, and an apparent decrease in the transport function of the anterior uvea. The extent and the duration of these effects were dose-dependent. The lower doses used, 3 or 6 micrograms of Cu as CuSO4 per eye, produced reversible inflammation. The highest dose, 30 micrograms of Cu per eye, also produced some signs of ocular chalcosis: hemorrhage, vitreous liquefaction, prolonged hypotony and local iridial ischemia. Six hours after the intravitreal injection of 6 micrograms of Cu as CuSO4 per eye, the Cu concentration in the vitreous humor increased to approximately 100 times that in the vitreous of control eyes, and began to decline only 3 days later, with a half-time of approximately 8 days. The Cu concentration in the anterior chamber of these eyes never exceeded 1 ppm and returned close to control values within 3 days. Based on these findings, factors that affect ocular trace-metal distribution and kinetics are discussed, as are reasons for the apparent difficulty in diagnosing the presence of Cu-containing intraocular foreign bodies on the basis of the Cu concentration of the aqueous humor.

Facilitation of copulatory performance in male rats by naloxone: effects of hypophysectomy, 17 alpha-estradiol, and luteinizing hormone releasing hormone.

Three experiments were performed to explore the mechanism whereby systemic administration of the opiate receptor antagonist, naloxone hydrochloride (20 mg/kg) causes reductions in the frequency of intromissions preceding ejaculation and latency to ejaculation in sexually experienced male rats. Administration of naloxone to male rats which were hypophysectomized in addition to being castrated and implanted SC with 30 mm silastic capsules containing testosterone caused such behavioral changes, suggesting that these behavioral effects of naloxone do not result from interference with the binding of endophin of pituitary origin. Surprisingly, a significant facilitatory effect of naloxone on sexual performance was absent in castrated controls bearing 30 mm testosterone implants. Recent evidence suggests that 17 alpha-hydroxylated estrogens, which may be produced in gonadally intact males, possess appreciable affinity for opioid receptors. However, daily administration of 17 alpha-estradiol (50 micrograms) to castrated, testosterone-implanted males failed to make them as behaviorally responsive to naloxone as gonadally intact animals. Administration of LHRH (1 microgram given SC 1.5 hr prior to testing) caused a significant reduction in ejaculation latency in gonadally intact males but not in castrated males bearing 30 mm testosterone implants. It is suggested that the facilitatory effect of naloxone on masculine sexual performance results, in part, from a drug-induced release of LHRH.

Facilitation by opiate antagonists of sexual performance in the male rat.

Intraperitoneal administration of increasing doses (5 mg/kg or 20 mg/kg) of the opiate antagonists naloxone hydrochloride or naltrexone hydrochloride to sexually experienced male rats caused a significant reduction in the number of intromissions prior to ejaculation and ejaculation latency. Mount and intromission latencies were significantly reduced following treatment with only the lower dose of each antagonist. Ways are suggested whereby endorphins may modulate masculine sexual performance and thereby contribute to successful fertilization of female conspecifics.

Evaluation of the BBL Minitek system for the identification of Enterobacteriaceae.

Thirty-one different substrate disks were tested in parallel with comparable, prepared media (BBL) against a minimum of 300 cultures of Enterobacteriaceae. An overall correlation of 98% was observed with all the disks tested. In addition, the system was used to identify 461 fresh isolates of Enterobacteriaceae in parallel with conventional media using the schema used at the Veterans Administration Hospital, Baltimore. An overall correlation of 97% was observed. Minitek is a time and space saving system. It is accurate and easily adapted to the clinical laboratory. A wide variety of substrates are available, allowing most laboratories to use their own schema. The long shelf life of most disks is a definite advantage.