A different perspective on sofosbuvir-ledipasvir treatment of patients with HCV genotype 1b cirrhosis: The ital-c network study.

The effectiveness of a 12-week course of sofosbuvir-ledipasvir in treatment-experienced HCV genotype 1b-infected patients with cirrhosis is still under debate. Our primary endpoint was to compare the sustained virological response at post-treatment week 12 (SVR12) of sofosbuvir-ledipasvir in combination with ribavirin for 12 weeks, and sofosbuvir-ledipasvir alone for 24 weeks. This was a prospective observational study that enrolled 424 (195 naive, 229 experienced; 164 treated for 12 weeks with Ribavirin and 260 with sofosbuvir-ledipasvir alone for 24 weeks) consecutive HCV genotype 1b-infected patients with cirrhosis. The SVR12 rates were 93.9% and 99.2% in patients treated for 12 and 24 weeks, respectively (P = .002). The baseline characteristics of patients treated for 12 weeks were significantly different from those treated for 24 weeks as regards their younger age (P = .002), prevalence of Child-Pugh class A (P = .002), lower MELD scores (P = .001) and smaller number of nonresponders (P = .04). The shorter treatment was significantly associated with a lower SVR12 in univariate and multivariate analyses (P = .007 and P = .008, respectively). The SVR rate was unaffected by age, gender, BMI, Child-Pugh class, MELD score or previous antiviral treatment. Patients receiving ribavirin experienced more episodes of ascites and headache but less recurrence of hepatocellular carcinoma (HCC), and were prescribed more diuretics and cardiopulmonary drugs. No patient discontinued treatment. The therapeutic regimen of sofosbuvir-ledipasvir plus ribavirin administered for 12 weeks was less effective than sofosbuvir-ledipasvir alone given for 24 weeks. At odds with European guidelines, the recommended 12-week treatment with sofosbuvir-ledipasvir alone might be suboptimal for this setting of patients.

Prospective observational study on the clinical behaviour of dental implants in patients with haemophilia. Preliminary results.

Haemophilia is a recessive congenital hereditary haemorrhagic disorder characterised by the decrease in, or absence of, the functional activity of factor VIII (Haemophilia A) or factor IX (Haemophilia B). The haematological medical treatment for these patients is systemic replacement therapy with factor VIII or factor IX concentrates. Dental implants are considered the gold standard for the replacement of missing teeth. There is no evidence or safe protocol for their use in patients with haemophilia. The objective of this study was to evaluate the clinical osseointegration and the incidence of postoperative bleeding in patients with haemophilia. The patients included in this study were over 18 years old with a diagnosis of haemophilia. A surgical implant protocol was implemented, supported by systemic and local haemostatic measures. Age, type of haemophilia, pre- and post-factor levels, gingival index, surgical duration, oedema, and pain were recorded for each case. In the event of haemorrhage, the bleeding index was determined. At four months, the success of the implant was evaluated using the absence of pain, suppuration, and clinical mobility as parameters. Fifteen surgeries were performed and 21 implants were placed in 10 patients. All the implants were successful in terms of the evaluated parameters. Among the 15 surgeries performed, bleeding was detected in three. The preliminary results found in this study seem to establish that the proposed surgical implant and haemostatic protocol is a predictable treatment for the placement of dental implants in patients with haemophilia.

Anosmia in the first coronavirus disease 2019 outbreak in Europe: functional recovery after eight months.

BACKGROUND: Severe acute respiratory syndrome coronavirus-2 is a formidable virus, responsible for coronavirus disease 2019 and endowed with marked neurotropism. The damage it causes to the nervous system is manifold. The main neurological manifestation is anosmia. Olfactory damage is often transient, but there are no data reflecting an observational period of several months. OBJECTIVE: This study evaluated the trend of anosmia in patients affected by coronavirus disease 2019 in the eight months following diagnosis. METHODS: Fifty-five subjects who presented with symptoms suggestive of coronavirus disease 2019 and who developed anosmia, between the end of February and the beginning of March 2020, were investigated. The patients were interviewed after eight months to determine functional recovery and assess the degree of recovery. RESULTS: Ninety-one per cent of the population reported olfactory recovery and, of these, 53 per cent had total recovery after eight months. Females and younger age groups seem slightly advantaged in functional recovery. The elderly population appears to have excellent prospects for full functional recovery. CONCLUSION: Anosmia represents a frequent neurological manifestation during coronavirus disease 2019. Fortunately, it is transient in most cases, and only a small percentage of patients affected by it report long-term functional deficits.

Ultrasound-guided percutaneous irreversible electroporation of hepatic and abdominal tumors not eligible for surgery or thermal ablation: a western report on safety and efficacy.

PURPOSE: To report our first results on sixteen patients affected by liver and abdominal malignant tumors, unfit for surgery or thermal ablation, treated with US-guided percutaneous irreversible electroporation (IRE). METHODS: From June 2014 to December 2016, all patients meeting the inclusion criteria (malignant hepatic or abdominal tumors not eligible for resection or thermal ablation) and not meeting the exclusion criteria (heart arrhythmia, pro-hemorrhagic hematological alterations, tumor size > 8 cm, presence of a biliary metallic stent) referred to our institutions were prospectively enrolled to undergo percutaneous US-guided irreversible electroporation (IRE). Sixteen patients (age range 59-68 years, mean 63; 7 females) with 18 tumors (diameter range 1.3-7.5 cm) fulfilled the inclusion criteria and were included in the study. Data concerning efficacy (tested by a 1-week CEUS and a 4-week enhanced CT and/or enhanced MRI) and safety were recorded during a 18-month follow up. RESULTS: All patients completed a 35-50-min procedure without complications. One patient with 6 cm Klatskin tumor also underwent a second session for 1 month. A 1-week CEUS and a 4-week e-CT and/or e-MRI arterial phase contrast enhancement analysis showed an overall reduction of arterial flow with confirmation of unenhanced lesions for seven nodules. After 1-18 months of follow up, no major complications were recorded and no tumor-related death occurred. The lesions of two patients disappeared 3 and 6 months after their treatment, respectively. CONCLUSIONS: IRE is a promising ablation modality in the treatment of malignant hepatic and abdominal tumors unsuitable for resection or thermal ablation.

Ectopic teeth in the maxillary sinus: A case report and literature review.

Ectopic eruption of teeth is a rare phenomenon although there have been reports of teeth in the nasal septum, mandibular condyle, and maxillary sinus. This impaction can present itself in a variety of ways such as chronic or recurrent sinusitis, sepsis, and facial numbness and can also be asymptomatic. The aim of this study was to describe, by means of research literature and by a case report, the characteristics and occurrence of ectopic eruption in the maxillary sinus. We have analyzed and compared clinical cases of ectopic teeth in the maxillary sinus with a search on PubMed utilizing keywords such as "ectopic," "teeth," "sinus," "maxillary," and Boolean operators "or" and "and" up until 2016. Fifty-one cases were found, of which 53% were female. The age ranged between 3 and 72 years, with an average age of 28.36 years. The higher prevalence of ectopic teeth is the 3rd molars. Ten of these teeth are associated with a dentigerous cyst, 1 by an osteoma, and 2 by soft tissue. Standard treatment for an ectopic tooth is extraction, but for other patients, treatment of choice in asymptomatic ectopic tooth cases is continued observation. Ectopic teeth tend to form a cyst or tumor if not managed.

Volatile compounds secreted by Brindley's glands of adult Triatoma infestans: identification and biological activity of previously unidentified compounds.

Volatile emissions of adult male Triatoma infestans were collected on non-polar SPME fibers and analyzed by gas chromatography linked to a mass spectrometer. A complex mixture of 16 short-chain esters and acids were identified. The composition of short-chain aliphatic acids (ethanoic to nonanoic acids) was similar to previously reported results. The most abundant aliphatic acid was 2-methylpropanoic acid, constituting 18% of the total volatile content. Also abundant were the esters 2- and 3-methylbutyl 2-methylpropanoate, which constituted 30% and 22%, respectively, of the total volatile content. A similar pattern of compounds was observed in the volatiles secreted by dissected male Brindley's glands; however, in this case, 2- and 3-methylbutan-1-ol were detected which were not found in live insect volatile emissions. Large variability in volatile composition was also observed among the glands excised from different insects. Electroantennographic (EAG) evaluation of the components of Brindley's gland showed significant responses for 2- and 3-methylbutyl 2-methylpropanoate compared to controls. The mixture of volatiles secreted by excised Brindley's glands and the isolated 2- and 3-methylbutyl 2-methylpropanoate had repellent effects on both male and female T. infestans, possibly associated with a defensive strategy.

Functional outcomes in patients reconstructed with flaps following surgery for hypopharyngeal cancer.

Following oncologic surgery for advanced cancer of the hypopharynx, primary closure of the defect of the upper aerodigestive tract is difficult to achieve. Usually locoregional or free flaps are used, the choice being determined by the extent of the surgical defect, the expertise of the surgeons and the general condition of the patient. Aim of the present study was to evaluate the functional recovery of patients who underwent surgical reconstruction, following hypopharyngeal cancer resection, with pedicled or free flaps. A retrospective analysis was conducted examining hospital records of the patients submitted to surgical treatment for hypopharyngeal cancer and reconstruction with pedicled or free flaps in the period between January 1995 and July 2004. Free flaps showed less severe complications, shorter hospital stay, less time to resume oral feeding compared with pedicled flaps. For this reason, we consider free flaps the gold standard for hypopharyngeal reconstruction, while pedicled flaps as the pectoralis major or other locoregional flaps should be used in those cases in which free flap reconstruction is not feasible or contraindicated.

Calcitonin-induced changes in the organization of sulfatide-containing membranes.

The interactions of salmon calcitonin with glycosphingolipid sulfatide are studied by right angle light scattering from the lipid suspension, by the excimer to monomer ratio (E/M) of the fluorescence intensity of pyrene phosphatidylcholine and pyrene sulfatide and by the leakage of carboxyfluorescein. It was found that calcitonin strongly modified the structure of the sulfatide aggregate, as indicated by the light scattering determinations. At a lipid peptide ratio 100:1 (molar ratio) light scattering from the suspension was negligible, indicating the formation of peptide-sulfatide complexes with a structure different from that of the lipid aggregate. The interactions of calcitonin with sulfatide when the latter is a component of a bilayer were also evaluated. A specific calcitonin-membrane sulfatide interaction was demonstrated by determining the temperature-dependent E/M of pyrene phosphatidylcholine and pyrene sulfatide in dipalmitoyl phosphatidylcholine/sulfatide (80:20, molar ratio) liposomes. The E/M curves were modified by calcitonin only when the liposomes were labelled with fluorescent sulfatide which probes the sulfatide behavior in the membrane. Furthermore, the addition of calcitonin to the incubation medium of liposomes containing sulfatide promoted the release of vesicle entrapped carboxyfluorescein without disrupting the bilayer structure, the release being correlated with the amount of sulfatide in the bilayer and the calcitonin concentration in the medium.

Plasma dependent pH sensitivity of liposomes containing sulfatide.

In this study we investigated the possibility to define relatively plasma-stable liposomal preparations in which the sensitivity to moderate drops of pH (i.e., from 7.4 to 6.8) would be induced by the presence of plasma itself. The liposome stability was monitored by determining the release of entrapped 5,6-carboxyfluorescein (CF). Using small unilamellar vesicles composed of egg phosphatidylcholine (EPC) and bovine brain sulfatide (CS) (4:1, molar ratio), the amount of CF released at pH 6.8 in the presence of 50% plasma was 3-fold that at pH 7.4, whereas no significant differences in the amount of CF released were observed when the same liposomes were incubated in buffer at pH 7.4 and 6.8, respectively. The increase in plasma induced leakage as a consequence of a drop in the pH medium, seems to specifically depend on the presence of sulfatide molecule in the bilayer since neither the acidic cholesterol 3-sulfate nor galactocerebroside, are able to induce pH sensitivity in EPC liposomes. Of all the plasma components considered (VLDL, LDL, HDL, protein fraction), VLDL seemed preferentially involved in the pH sensitivity induced by CS since they promoted an almost complete release of CF from EPC/CS small unilamellar vesicles. Thus, these liposomes are potentially a useful tool for a specific drug delivery to those pathological tissues such as tumors, inflammation sites and ischemic areas in which it is known that a lowering of the pH can occur.

Pyrene lipids as markers of peroxidative processes in different regions of low and high density lipoproteins.

Three different pyrene derivatives, pyrene decanoyl phosphatidylcholine (P10PC), pyrene dodecanoyl sulfatide (P12CS) and cholesteryl pyrenyl hexanoate (P6Chol), were used to follow lipid peroxidation in low and high density lipoproteins. Probe-labelled lipoproteins were subjected to Cu2+ catalyzed peroxidation. In all cases the fluorescence of the probes progressively decreased due to the involvement of pyrene in the peroxidative reaction. Thus, we used the fluorescence decrease of P6Chol to monitor the lipid peroxidation in the hydrophobic core of LDL and HDL, and that of the amphipatic probes, P10PC and P12CS, to follow lipid peroxidation in the envelope of both lipoproteins. The possibility of following lipid peroxidation in individual lipoprotein regions could lead to more detailed information on the oxidative modifications that play an important role in the altered cholesterol homeostasis involved in the formation of atherosclerotic lesions. No differences were observed in the peroxidation kinetics of the hydrophobic core of HDL and LDL monitored with P6Chol. On the contrary kinetics obtained with P10PC and P12 CS demonstrated the HDL envelope to be more susceptible to Cu2+ -dependent lipid peroxidation than that of the LDL. This could be due to a greater radical generating capacity of the HDL envelope and can be explained on the basis of low vitamin E levels and large amounts of polyunsaturated fatty acids esterified on phospholipids determined in HDL, and on literature evidence that indicates HDL as the principal vehicle of circulating plasma lipids peroxides.

HCV genotype 1 subtypes (1a and 1b): similarities and differences in clinical features and therapeutic outcome.

AIM: To evaluate similarities and differences in HCV-1 subtypes 1a and 1b in the presenting clinical features and the response to peg-interferon and ribavirin (Peg/RIBA). PATIENTS AND METHODS: A total of 1,233 naïve patients with HCV genotype-1 infection, 159 (13%) with subtype 1a and 1,074 (87%) with subtype 1b were treated with Peg-IFN/RIBA at 12 Italian centers. Covariates included in the logistic model were age, gender, BMI, serum alanine aminotransferase, serum gamma-glutamiltranspeptidase (γGT), platelets counts, liver fibrosis, the occurrence of type 2 diabetes, baseline viremia, and IL28B genotype. RESULTS: At multivariate analysis, baseline characteristics differentiating patients with HCV-1a versus HCV-1b were young age, male gender, no F4 fibrosis, and no diabetes. SVR was achieved by 37% of patients with subtype 1b and 45% of those with subtype 1a, a nonsignificant difference of 8% (p = 0.069). In patients with subtype 1a, predictors of SVR were IL28B CC (OR 5.78, CI 1.98-16.83), RVR (OR 4.18, CI 1.66-10.55), female gender (OR 2.83, CI 1.83-6.78), and HCVRNA (OR 0.55, CI 0.32-0.96). In patients with subtype 1b, the ranking of predictors was levels RVR (OR 6.49, CI 4.32-9.73), IL28B CC (OR 3.32, CI 2.15-4.58), γGT (OR 1.59, CI 0.14-2.22), HCVRNA (OR 0.61, CI 0.47-0.79), and age (OR 0.01, CI 0.02-0.42). CONCLUSION: In Italy HCV-1 subtype 1a prevails in young male patients with less advanced liver damage, findings that imply a more recent spreading of the infection with this viral strain. The two HCV-1 subtypes appear equally responsive to Peg-IFN/RIBA, with IL28B genotyping and monitoring of RVR mostly influencing the therapeutic response.

Characterization of reactive oxygen species induced effects on human spermatozoa movement and energy metabolism.

Reactive oxygen species (ROS) inhibit sperm movement and have been implicated in male infertility. In this study, we determined the effects of specific ROS produced by activated leukocytes on human spermatozoa and investigated their metabolic site of action. We used chemiluminescence and electron paramagnetic resonance (EPR) to characterize the ROS generated by both blood and seminal leukocytes. We also determined the effects of these ROS on sperm energy metabolism using biochemical analyses and flow cytometry. Both blood and seminal leukocytes produced the same characteristic ROS which were determined to be hydrogen peroxide (H2O2) and superoxide radicals (O2*-). EPR using the spin trapping technique indicated that superoxide radical-dependent hydroxyl radicals (HO.) were also generated. ROS generated by PMA-stimulated blood leukocytes (2-5 x 10(6)/ml) caused inhibition of sperm movement in 2 h (p < .01). Using the hypoxanthine/ xanthine oxidase (0.5 U/ml) system to generate ROS, we determined that spermatozoa ATP levels, after ROS treatment, were reduced approximately eight-fold in 30 min (0.10 x 10(10) moles/10(6) sperm cells) compared to control (0.84 X 10(-10) moles/10(6) sperm cells) (p < .01). Sperm ATP reduction paralleled the inhibition of sperm forward progression. Neither superoxide dismutase (100 U/ml) nor dimethyl sulfoxide (100 mM) reversed these effects; however, protection was observed with catalase (4 X 10(3) U/ml). Flow cytometric analyses of sperm treated with various doses of H2O2 (0.3 mM-20.0 mM) showed a dose-dependent decrease in sperm mitochondrial membrane potential (MMP); however, at low concentrations of H2O2, sperm MMP was not significantly inhibited. Also, sperm MMP uncoupling with CCClP had no effect on either sperm ATP levels or forward progression. These results indicate that H2O2 is the toxic ROS produced by activated leukocytes causing the inhibition of both sperm movement and ATP production. O2*- and HO. do not play a significant role in these processes. Low concentrations of H2O2 causing complete inhibition of sperm movement and ATP levels inhibit sperm energy metabolism at a site independent of mitochondrial oxidative phosphorylation.

Adrenaline-synthesizing neurons in the medulla of the cat.

In this study, the distribution of neurons containing the adrenaline-synthesizing enzyme phenylethanolamine-N-methyltransferase (PNMT) was mapped in the medulla of the cat. Data from recent studies in the rat suggest that the anatomical structure responsible for cardiorespiratory changes that occur following application of neurotransmitters and drugs to Schlaefke's area on the ventral medullary surface is the nucleus reticularis rostroventrolateralis (RVL), which is distinguished from adjacent regions of the reticular formation, in part, by the presence of adrenaline-synthesizing neurons. To determine whether an equivalent adrenergic population is present in the RVL of the cat, we used antibodies raised against bovine adrenal PNMT to map the distribution of adrenaline-synthesizing neurons in the reticular formation. In the ventrolateral medulla, we found that labeled cells extended from the level of the retrofacial nucleus to the calamus scriptorius. The majority of labeled cells were seen in a nucleus designated RVL at the level of the rostral one-third of the inferior olive. In the dorsomedial medulla, cells were labeled in the caudal aspect of the nucleus tractus solitarii (NTS) and were especially dense in the subnucleus gelatinosus and commissural nucleus of the vagus. A few lightly labeled cells were also present in the rostral pole of the area postrema (AP). In contrast to the rat, few or no immunoreactive cells were found in the rostral NTS, medial longitudinal fasciculus, nucleus paragigantocellularis dorsalis, or periventricular gray. Our results are consistent with the notion that an area of the RVL containing adrenergic perikarya is the anatomical structure responsible for cardiovascular changes that occur when chemicals are applied to Schlaefke's area.

Effects of two oral doses of alendronate in the treatment of Paget's disease of bone.

Twenty patients with mild Paget's disease of bone were given either 20 (10 patients) or 40 mg alendronate daily for 6 months. The 20-mg dose was well tolerated, but in 3 patients on 40 mg/d alendronate, the treatment was withdrawn after 3-5 months because of gastric and oesophageal disturbances. Urinary hydroxyproline excretion fell within the first month to 77 +/- 5% (SD) and to 47 +/- 5% of pretreatment values in the 20- and 40-mg dosing group, respectively (p < 0.001 between group comparison). The serum alkaline phosphatase fell more slowly with the maximum suppression of disease activity reached at 4 months, when it attained a plateau in both groups of patients. However, the decrease in serum alkaline phosphatase was significantly more pronounced in the patients treated with 40 mg/d tablets (50 +/- 10% of pretreatment values) than in those given 20 mg alendronate per day (76 +/- 9% of initial value), in none of whom a disease remission was observed. It appears, therefore, that while 20 mg/d oral doses of alendronate are insufficient, 40 mg/d are associated with a high incidence of side effects. Furthermore, the suppression of disease activity depends on the dose of bisphosphonate given daily or over a short period of time and lower doses cannot be compensated by a longer duration of the treatment course.

The effect of a vasopressin antagonist on the pressor response to histamine administered centrally.

The mechanism for the pressor response to intracerebroventricularly (i.c.v.) administered histamine was studied. Histamine (HA), when injected intracerebroventricularly in rats, produced a dose-dependent increase in mean arterial pressure (MAP) which was subject to tachyphylaxis. Spinal transection at C-7 in the anesthetized rat did not attenuate the rise in blood pressure. The possibility that the release of 8-arginine vasopressin was responsible for the rise in blood pressure was explored. By pretreating conscious freely-moving rats with a specific antagonist to the vasopressor action of vasopressin viz., [1-beta-mercapto-beta, beta-cyclopentamethyleneproprionic acid), 2-(O-methyl) tyrosine] arginine-vasopressin, there was a statistically-significant attenuation of the pressor response to intracerebroventricularly administered histamine. The antagonist however, did not totally abolish the pressor response, regardless of the dose employed. Concomitant administration of hexamethonium and the vasopressin antagonist did not further attenuate the response. Previous adrenal demedullation did not diminish the response to intracerebroventricularly administered histamine, nor was there any evidence for release of angiotensin II since pretreatment with saralasin did not attenuate the cardiovascular response. These findings suggest that vasopressin along with other as yet undefined substances, are released from the central nervous system to produce the increase in blood pressure after intracerebroventricularly administered histamine.

The effect of intrahypothalamic injection of homodimaprit on blood pressure.

Bilateral injection of the inhibitor of histamine-N-methyltransferase, SKF 91488, which is also known as homodimaprit (5 micrograms), into the preoptic area of the rat produced delayed hypertension, tachycardia and hyperthermia. Some animals exhibited pulmonary edema. These effects were only noted 18-24 hr after an injection and were not an artifact of the injection, since the administration of artificial cerebrospinal fluid produced none of these effects. At the time noted, lesions of the rostral hypothalamus, including the preoptic area, were evident. Injection of a vasopressin antagonist, intravenously, did not lower the blood pressure of the hypertensive animals nor did previous bilateral adrenal demullation prevent or delay the hypertension or tachycardia. Therefore, it does not appear that hypersecretion of either vasopressin or adrenal catecholamines contributed to the cardiovascular effects. Peripheral pretreatment with the sympathetic neurotoxin 6-hydroxydopamine however, did prevent the delayed rise in blood pressure following an injection of homodimaprit. From these studies, it is concluded that the injection of homodimaprit produces lesions in the preoptic area, resulting in hypertension that is maintained by excessive activation of the sympathetic nervous system.

Cardiorespiratory effects produced by injecting drugs that affect GABA receptors into nuclei associated with the ventral surface of the medulla.

It has recently been shown that L-glutamic acid induced stimulation of cell bodies in a circumscribed area of the rostral ventrolateral medulla (RVLM) in the cat, produced increases in arterial pressure (AP), decreases in heart rate (HR) and transient apnea (Gatti, Norman, DaSilva and Gillis, 1986). The purpose of the present study was to determine if this same area was sensitive to GABA receptor agonists and antagonists. Injection of the GABA agonist muscimol (200 ng), into the rostral ventrolateral medulla of cats anesthetized with chloralose produced a precipitous and immediate fall in arterial pressure (-95 +/- 4.6) and heart rate (-31 +/- 5.9; n = 4, P less than 0.05). Maximal cardiovascular effects could only be achieved if muscimol was injected bilaterally. These effects of muscimol on arterial pressure were dose-dependent. Time-action curves for the effects of muscimol on arterial pressure and respiration were different. Hypotension occurred first and was followed later in time by a decrease in minute ventilation. Within 30 min all animals were apneic after the 200 ng dose. The cardiovascular effects of muscimol were reversed by the injection of the GABA receptor antagonist bicuculline. These data indicate that stimulation of GABA receptors in the rostral ventrolateral medulla produced selective cardiovascular effects and that respiratory neurons sensitive to GABA are apparently not localized with these cardiovascular neurons.

Synthesis and structure-activity relationships of bafilomycin A1 derivatives as inhibitors of vacuolar H+-ATPase.

The macrolide antibiotic bafilomycin A1 is a highly potent and selective inhibitor of all the vacuolar ATPases (V-ATPases). With the aim of obtaining novel analogues specific for the osteoclast subclass of vacuolar ATPase, 31 derivatives of bafilomycin A1 were synthesized and tested for their ability to inhibit differentially the V-ATPase-driven proton transport in membrane vesicles derived from chicken osteoclasts (cOc) and bovine chromaffin granules (bCG). Although none of the new analogues were more potent than the parent compound, the obtained data provided a significant amount of information about the structural requirements for the inhibitory activity of bafilomycin A1. The different effects of a few analogues on the two enzymes could also suggest the possibility of a selective modulation of the V-ATPases in different tissues.

Inhibition of HIV type 1 production by hygromycin B.

HIV infection alters the cellular uptake of ions and other small molecules. This study was designed to determine whether hygromycin B, a low molecular weight (MW 527) aminoglycoside protein synthesis inhibitor that is normally impermeable to mammalian cells at micromolar concentrations, can selectively inhibit HIV expression and cytopathology. CD4+ T lymphoblastoid cells (H9) and peripheral blood mononuclear cells (PBMCs) were infected with HIV-1, then incubated in medium containing various concentrations of hygromycin B. HIV-1-induced formation of multinucleated giant cells and single cell killing were dramatically reduced in the presence of micromolar concentrations of hygromycin B. Hygromycin B also inhibited HIV-1 production in a dose-dependent manner during acute infection. G418, a larger and more hydrophobic aminoglycoside (MW 692), did not display the same selective inhibition of HIV-1 production as hygromycin B. Relative to mock-infected cells, protein synthesis in acutely infected H9 cells was selectively inhibited by hygromycin B. Hygromycin B also reduced HIV production in PBMCs and in H9 cells persistently infected with HIV. PCR analysis demonstrated that hygromycin B did not inhibit HIV-1 reverse transcription. These results demonstrate that HIV-1 infection renders cells more sensitive to hygromycin B than uninfected cells, and provides support for the hypothesis that HIV-1 induces an alteration of plasma membrane permeability. The HIV-modified cell membrane may be a potential target for antiviral intervention and chemotherapy.

A synthetic peptide corresponding to the carboxy terminus of human immunodeficiency virus type 1 transmembrane glycoprotein induces alterations in the ionic permeability of Xenopus laevis oocytes.

The carboxy-terminal 29 amino acids of the human immunodeficiency virus type 1 transmembrane glycoprotein (HIV-1 TM) are referred to as lentivirus lytic peptide 1 (LLP-1). Synthetic peptides corresponding to LLP-1 have been shown to induce cytolysis and to alter the permeability of cultured cells to various small molecules. To address the mechanisms by which LLP-1 induces cytolysis and membrane permeability changes, various concentrations of LLP-1 were incubated with Xenopus laevis oocytes, and two-electrode, voltage-clamp recording measurements were performed. LLP-1 at concentrations of 75 nM and above induced dramatic alterations in the resting membrane potential and ionic permeability of Xenopus oocytes. These concentrations of LLP-1 appeared to induce a major disruption of plasma membrane electrophysiological integrity. In contrast, concentrations of LLP-1 of 20-50 nM induced changes in membrane ionic permeability that mimic changes induced by compounds, such as the bee venom peptide melittin, that are known to form channel-like structures in biological membranes at sublytic concentrations. An analog of LLP-1 with greatly reduced cytolytic activity failed to alter the electrophysiological properties of Xenopus oocytes. Thus, by altering plasma membrane ionic permeability, the carboxy terminus of TM may contribute to cytolysis of HIV-1-infected CD4+ cells.