Mindfulness Meditation Programs Informed by Transgender Youth.

Objectives: This study explores the openness of transgender and gender diverse youth and young adults (TGDY) to mindfulness meditation programs in order to create culturally informed interventions to benefit this population. Method: Two focus groups were conducted with a total of ten TGDY ages 14-24 years old at a transgender youth health center in a large metropolitan city in the USA. A 10-min guided mindfulness meditation was included for participants to experience and voice reactions to. The State-Trait Anxiety Inventory (STAI) was utilized to measure the quantitative impact of the meditation on participants' anxiety and thematic analysis for the qualitative data. Results: Reflexive Thematic Analysis on qualitative focus group data revealed the following four themes: Active in Self-care, Silent Meditation Is "Not for Me," Guided Mindfulness Calms and Connects, and Program Ideas for Future. STAI results indicated a statistically significant reduction in anxiety following participation in the group meditation. Conclusions: Participants were open to mindfulness as an additional method of self-care, and they emphasized future programs should include sensory stimulation, a pressure-free environment accepting of active minds and bodies, and a transgender instructor if possible. Meditation and mindfulness have the potential to be a very powerful healing modality for TGDY in clinical and therapeutic care. Preregistration: This study is not preregistered. Supplementary Information: The online version contains supplementary material available at 10.1007/s12671-022-02048-6.

The structures of penicillin-binding protein 4 (PBP4) and PBP5 from Enterococci provide structural insights into ß-lactam resistance.

The final steps of cell-wall biosynthesis in bacteria are carried out by penicillin-binding proteins (PBPs), whose transpeptidase domains form the cross-links in peptidoglycan chains that define the bacterial cell wall. These enzymes are the targets of ß-lactam antibiotics, as their inhibition reduces the structural integrity of the cell wall. Bacterial resistance to antibiotics is a rapidly growing concern; however, the structural underpinnings of PBP-derived antibiotic resistance are poorly understood. PBP4 and PBP5 are low-affinity, class B transpeptidases that confer antibiotic resistance to Enterococcus faecalis and Enterococcus faecium, respectively. Here, we report the crystal structures of PBP4 (1.8 Å) and PBP5 (2.7 Å) in their apo and acyl-enzyme complexes with the ß-lactams benzylpenicillin, imipenem, and ceftaroline. We found that, although these three ß-lactams adopt geometries similar to those observed in other class B PBP structures, there are small, but significant, differences that likely decrease antibiotic efficacy. Further, we also discovered that the N-terminal domain extensions in this class of PBPs undergo large rigid-body rotations without impacting the structure of the catalytic transpeptidase domain. Together, our findings are defining the subtle functional and structural differences in the Enterococcus PBPs that allow them to support transpeptidase activity while also conferring bacterial resistance to antibiotics that function as substrate mimics.

Overcoming multidrug resistance using folate receptor-targeted and pH-responsive polymeric nanogels containing covalently entrapped doxorubicin.

Multidrug resistance (MDR) contributes to failure of chemotherapy. We here show that biodegradable polymeric nanogels are able to overcome MDR via folic acid targeting. The nanogels are based on hydroxyethyl methacrylamide-oligoglycolates-derivatized poly(hydroxyethyl methacrylamide-co-N-(2-azidoethyl)methacrylamide) (p(HEMAm-co-AzEMAm)-Gly-HEMAm), covalently loaded with the chemotherapeutic drug doxorubicin (DOX) and subsequently decorated with a folic acid-PEG conjugate via copper-free click chemistry. pH-Responsive drug release is achieved via the acid-labile hydrazone bond between DOX and the methacrylamide polymeric network. Cellular uptake and cytotoxicity analyses in folate receptor-positive B16F10 melanoma versus folate receptor-negative A549 lung carcinoma cells confirmed specific uptake of the targeted nanogels. Confocal microscopy demonstrated efficient internalization, lysosomal trafficking, drug release and nuclear localization of DOX. We also show that DOX resistance in 4T1 breast cancer cells results in upregulation of the folate receptor, and that folic acid targeted nanogels can be employed to bypass drug efflux pumps, resulting in highly efficient killing of resistant cancer cells. In conclusion, folic acid functionalized nanogels with pH-controlled drug release seem to hold significant potential for treating multidrug resistant malignancies.

A meta-analysis of efficacy and safety of catheter-directed interventions in submassive pulmonary embolism.

OBJECTIVE: Catheter-directed interventions, such as catheter-directed thrombolysis (CDT), are becoming a popular therapeutic option for patients with hemodynamically stable pulmonary embolism (PE) and right ventricle (RV) dysfunction (submassive PE). We wished to quantitatively assess therapeutic efficacy and safety of catheter-directed interventions in submassive PE. MATERIALS AND METHODS: PubMed, Embase, Cochrane and Scopus were searched for studies on catheter-directed interventions and submassive PE. Studies reporting data on therapeutic efficacy (RV to left ventricle [RV/LV] ratio, systolic pulmonary artery pressure) and safety outcomes (in-hospital and 30-day mortality rates, major and minor bleeding rates) were retained and assessed. RESULTS: The final reference sample included 13 publications (11 papers and 2 conference abstracts), collectively enrolling 422 patients with submassive PE. The majority (8/13) studies were retrospective studies. One study was a randomized controlled study. Nine of 13 studies utilized CDT with or without ultrasound-assisted thrombolysis. The post-therapy pooled mean change of RV/LV ratio was -0.3 (95% confidence interval [CI]: -0.42, -0.18), and the pooled mean decrease of pulmonary artery pressure was -19.41 (95% CI: -27.65, -11.17) mm Hg. Safety outcome analysis demonstrated low pooled rates of adverse events (in-hospital mortality: 0.00 [95% CI: 0.00, 0.01]; 30-day mortality: 0.00 [95% CI: 0.00, 0.03]; major bleeding: 0.00 [95% CI: 0.00, 0.02]; minor bleeding: 0.05 [95% CI: 0.01, 0.12]). CONCLUSIONS: This meta-analysis demonstrates evidence of therapeutic efficacy and low rates of adverse events of catheter-directed interventions in submassive PE.

PEG stabilized DNA - poly(ferrocenylsilane) polyplexes for gene delivery.

Polycationic poly(ferrocenylsilane)s (PFS) with tunable amounts of PEG side chains were used for the condensation of DNA into polyplexes of 110 nm in 5.0 mM HEPES. The PFS-PEG/DNA polyplexes showed negligible aggregation, a strongly reduced protein adsorption, transfection activities comparable with linear polyethyleneimine and an excellent cytocompatibility.

Tea polyphenols inactivate Cronobacter sakazakii isolated from powdered infant formula.

This study evaluated the antimicrobial activity of tea polyphenols (TP) against 4 Cronobacter sakazakii strains with different sequence types (ST) isolated from powdered infant formula (PIF). The results showed that in normal saline, 5mg/mL of TP (pH 3.44) could eliminate approximately 7.0 log cfu/mL of C. sakazakii within 1 h; in rehydrated PIF, after acidification with HCl (pH 3.55), TP showed a stronger antibacterial activity compared with the controls (malic acid, ascorbic acid, and citric acid). Further, some differences were obvious in tolerance to TP between C. sakazakii strains with different ST. The tolerance of C. sakazakii CE1 (ST4) to TP was found to be greater than that of the other 3 C. sakazakii strains (ST1, ST8, and ST64). The results of recovered test and transmission electron microscope analysis revealed that the action of TP against C. sakazakii was an irreversible bactericidal process caused by leakage of cytoplasm. Taken together, these results indicated that TP had an effective bactericidal effect against C. sakazakii, and provided a new idea for preventing and inactivating C. sakazakii in PIF.

Correlation of 68Ga Ventilation-Perfusion PET/CT with Pulmonary Function Test Indices for Assessing Lung Function.

UNLABELLED: Pulmonary function tests (PFTs) are routinely used to assess lung function, but they do not provide information about regional pulmonary dysfunction. We aimed to assess correlation of quantitative ventilation-perfusion (V/Q) PET/CT with PFT indices. METHODS: Thirty patients underwent V/Q PET/CT and PFT. Respiration-gated images were acquired after inhalation of (68)Ga-carbon nanoparticles and administration of (68)Ga-macroaggregated albumin. Functional volumes were calculated by dividing the volume of normal ventilated and perfused (%NVQ), unmatched and matched defects by the total lung volume. These functional volumes were correlated with forced expiratory volume in 1 s (FEV1), forced vital capacity (FVC), FEV1/FVC, and diffusing capacity for carbon monoxide (DLCO). RESULTS: All functional volumes were significantly different in patients with chronic obstructive pulmonary disease (P < 0.05). FEV1/FVC and %NVQ had the highest correlation (r = 0.82). FEV1 was also best correlated with %NVQ (r = 0.64). DLCO was best correlated with the volume of unmatched defects (r = -0.55). Considering %NVQ only, a cutoff value of 90% correctly categorized 28 of 30 patients with or without significant pulmonary function impairment. CONCLUSION: Our study demonstrates strong correlations between V/Q PET/CT functional volumes and PFT parameters. Because V/Q PET/CT is able to assess regional lung function, these data support the feasibility of its use in radiation therapy and preoperative planning and assessing pulmonary dysfunction in a variety of respiratory diseases.

Serotypes, genotypes and antimicrobial resistance patterns of human diarrhoeagenic Escherichia coli isolates circulating in southeastern China.

Diarrhoeagenic Escherichia coli (DEC) infection is a major health problem in developing countries. The prevalence and characteristics of DEC have not been thoroughly investigated in China. Consecutive faecal specimens from outpatients with acute diarrhoea in nine sentinel hospitals in southeastern China were collected from July 2009 to June 2011. Bacterial and viral pathogens were detected by culture and RT-PCR, respectively. DEC isolates were further classified into five pathotypes using multiplex PCR. The O/H serotypes, sequence types (STs) and antimicrobial susceptibility profiles of the DEC isolates were determined. A total of 2466 faecal specimens were collected, from which 347 (14.1%) DEC isolates were isolated. DEC was the dominant bacterial pathogen detected. The DEC isolates included 217 EAEC, 62 ETEC, 52 EPEC, 14 STEC, one EIEC and one EAEC/ETEC. O45 (6.6%) was the predominant serotype. Genotypic analysis revealed that the major genotype was ST complex 10 (87, 25.6%). Isolates belonging to the serogroups or genotypes of O6, O25, O159, ST48, ST218, ST94 and ST1491 were highly susceptible to the majority of antimicrobials. In contrast, isolates belonging to O45, O15, O1, O169, ST38, ST226, ST69, ST31, ST93, ST394 and ST648 were highly resistant to the majority of antimicrobials. DEC accounted for the majority of bacterial pathogens causing acute diarrhoea in southeastern China, and it is therefore necessary to test for all DEC, not only the EHEC O157:H7. Some serogroups or genotypes of DEC were highly resistant to the majority of antimicrobials. DEC surveillance should be emphasized.

Isolation and characterization of novel microsatellite loci in Nibea albiflora.

Eleven novel microsatellite loci were isolated from a (CA)10-enriched genomic DNA library of Nibea albiflora. The characteristics of these microsatellites were determined in a sample of 48 N. albiflora individuals. The number of alleles at the 11 microsatellite loci ranged from 5 to 25, with an average of 13.5 per locus. The observed and expected heterozygosities varied from 0.583 to 0.917 and from 0.568 to 0.964, respectively. Eight of the 11 microsatellite loci conformed to the Hardy-Weinberg equilibrium. No significant linkage disequilibrium was found among all 11 loci. These polymorphic microsatellites will be useful for population genetic analyses of N. albiflora.

Development and characterization of novel microsatellite markers in the rock bream fish Oplegnathus fasciatus.

The rock bream fish Oplegnathus fasciatus is one of the most popular aquaculture species in China. In the present study, 15 novel polymorphic microsatellite loci were isolated and characterized from a wild population of O. fasciatus from the Zhoushan coast of China. The number of alleles per polymorphic locus ranged from 4 to 9 in a sample of 30 individuals. Observed and expected heterozygosities per locus varied from 0.267 to 0.767 and from 0.395 to 0.859, respectively. Eleven of the 15 microsatellite loci conformed to Hardy-Weinberg equilibrium. No significant linkage disequilibrium between pairs of loci was detected. The present microsatellite markers could provide a useful tool for the genetic analyses of O. fasciatus.

Biological control of postharvest sour rot of citrus by two antagonistic yeasts.

AIMS: To investigate antifungal effects of two antagonistic yeasts on postharvest sour rot caused by Geotrichum citri-aurantii in citrus and evaluate possible mechanisms. METHODS AND RESULTS: Cell suspension of Cryptococcus laurentii at 10(8) to 10(9) cells per ml effectively reduced sour rot incidence from 55.6% among untreated control fruit to 29.9-20.7% after 5 days of incubation at 26 degrees C. Application of cell-free culture filtrate of C. laurentii was effective in reducing the sour rot, but the effectiveness was lower than that of the cell suspension (1 x 10(8) cells per ml). In addition, C. laurentii multiplied more rapidly than Rhodosporidium paludigenum at 26 degrees C. The fruit inoculated with the two yeasts demonstrated changes in peroxidase and superoxide dismutase activity. Cryptococcus laurentii, in particular, was capable of inducing a striking response in treated citrus fruits. CONCLUSIONS: The antifungal ability of C. laurentii can be attributed to the competitions of nutrients and space, defensive responses and possible secretion of antibiotic compounds. SIGNIFICANCE AND IMPACT OF THE STUDY: The use of such antagonists may constitute an important alternative to synthetic fungicides for controlling postharvest sour rot in citrus.

First Report of Leaf Blight on Duying Caused by Phyllosticta anacardiacearum in China.

Duying (Elaeocarpus glabripetalus Merr.; Elaeocarpaceae) is widely cultivated as an ornamental tree of commercial importance in southern China. From 2003 to 2008, severe outbreaks of Duying leaf blight occurred in the Hangzhou area, Zhejiang Province. Disease incidence was greater than 20% and mainly infected young leaves and shoots in the spring and autumn. Severely infected leaves and shoots died and eventually led to branch dieback. The overall growth decline of affected trees occurs over 4 to 6 years before tree death. Infection symptoms are characterized by grayish, round, semicircular- or irregular-shaped spots (5 mm to 5 cm long) with dark brown borders and the appearance of black, granular pycnidia within the dead leaf tissues. The primary infection zones are commonly observed on the leaf margins and apices, are brown, up to 2 mm in diameter, and often surrounded by a yellow zone. Pycnidia were globose and 122 to 127 µm (average 123.5 µm) in diameter. A fungus was consistently isolated from symptomatic tissues on potato dextrose agar (PDA). Ash-black pycnidia appeared on PDA after 10 days. Ascospores developed on modified PDA (1 liter of PDA + 20 g of Duying leaves) after 18 days. Conidiogenous cells were cylindrical to obpyriform. The hyaline conidia were obovoid and guttulate, 10 to 13 × 6 to 8 µm (average 11.5 × 7.5 µm), and usually surrounded by a mucilaginous sheath with a hyaline apical appendage that was 5 to 8 µm long. Pseudothecia were solitary and subglobose with long necks. Asci were 45 to 70 × 7.5 to 12 µm (average 62.5 × 10.8 µm). Ascospores were 12 to 13 × 4 to 5 µm with rounded apices and hyaline, mucilaginous, apical caps. The fungus was morphologically identified as Phyllosticta anacardiacearum van der Aa (teleomorph Guignardia mangiferae A. J. Roy). This identification was also confirmed by the China General Microbiological Culture Collection Center (CGMCC). Six representative fungal isolates were identified by sequencing the internal transcribed spacer (ITS) region of the rDNA and comparing the sequences with those in GenBank using BLAST searches. The ITS sequences of six cultures (GenBank Accession Nos. EU821356-EU821361) showed 100% identity with the ITS sequences of an isolate of a Phyllosticta sp. (GenBank Accession No. AF532314) (2) and G. mangiferae (GenBank Accession No. AY277717) (1). To fulfill Koch's postulates, a conidial suspension (106 conidia per ml) collected from PDA cultures (isolate phy01) was used to spray inoculate leaves of potted 3-year-old Duying trees. Inoculated trees were kept for 48 h under a polyethylene sheet cover and grown at 10 to 15°C in a greenhouse. A total of 30 leaves of five healthy trees were inoculated with the pathogen. In addition, five 3-year-old trees were sprayed with sterile water to serve as uninoculated controls. After 10 to 14 days, inoculated leaves showed infection symptoms resembling those observed on Duying trees naturally infected with P. anacardiacearum. The pathogen was reisolated from the margins of necrotic tissues, but not from controls. To our knowledge, this is the first report of leaf blight on E. glabripetalus caused by P. anacardiacearum in China. Reference: (1) F. R. Katia et al. Mycol. Res. 108:45, 2004. (2) A. K. Pandey et al. Mycol. Res. 107:439, 2003.

First Report of Blackleg of Ajuga multiflora Caused by Phoma multirostrata in China.

Ajuga multiflora Bunge (Labiatae) is an ornamental that is widely planted in gardens in southern China. Blackleg symptoms were observed on A. multiflora during the autumn of 2005 in the Hangzhou area, Zhejiang Province. A. multiflora had to be replaced by other landscape plants during the past 2 years because of a serious outbreak of the disease. When the plant is in continuous cultivation for more than 2 years, disease incidence is more than 80%. Brown water-soaked spots appeared initially on the base of the petioles. Blackleg lesions from multiple infections may coalesce. Lesions often expand rapidly along the petioles followed by wilting and drying of individual leaves, ultimately resulting in the death of plants. The disease can occur at any time of year, but is most severe during the flowering stage. A fungus was consistently isolated from symptomatic tissues on potato dextrose agar. After 14 days on SNA (synthetic low nutrient agar; 1.0 g of KH2PO4, 1.0 g of KNO3, 0.5 g of MgSO4·7H2O, 0.5 g of KCl, 0.2 g of glucose, 0.2 g of sucrose, and 20 g of agar per liter) medium, the fungus produced light olive-to-gray mycelium, and later, generated brown, globose to subglobose pycnidia with hyaline, unicellular, and ellipsoidal conidia. Diameter of pycnidia ranged from 163 to 260 µm (average 203 µm). Conidiophores measured 3.5 to 6.2 × 2.0 to 3.2 µm (average 5.4 × 2.3 µm). The fungus was identified as Phoma multirostrata (P.N. Mathur, S.K. Menon & Thirum) Dorenb. & Boerema based on its morphological characteristics (1). The fungus grew between 4 and 36°C and optimum growth was at 23°C. To fulfill Koch's postulates, 15 healthy 20-day-old A. multiflora seedlings were planted in sterile soil artificially infested with P. multirostrata, which was grown on a mixture (1:10 w/w) of corn meal and sand. Soil was amended with 20% (w/w) of inoculum mixture. Another 15 plants grown in noninfested soil served as controls. All plants were grown at 22 to 25°C in the greenhouse. After 5 to 7 days, inoculated plants showed symptoms closely resembling those seen on plants from the garden. P. multirostrata was isolated from the margins of necrotic tissues on diseased plants, but not from the control plants. To our knowledge, this is the first report of P. multirostrata causing blackleg of A. multiflora in China. Reference: (1) B. C. Sutton. The Coelomycetes. CMI, Kew, Surrey, UK, 1980.

The interactions of phenytoin and its binding site in DI-S6 segment of Na+ channel voltage-gated peptide by NMR spectroscopy and molecular modeling study.

Nuclear magnetic resonance (NMR) spectra of a model peptide (BL-DIS6), in the presence of anticonvulsant diphenyl drug, phenytoin (DPH), were measured to obtain the interactions between the selected drug and the model peptide. BL-DIS6's sequence corresponds to the S6 segment in domain I of rat brain type IIA Na+-channel. NMR studies have demonstrated that the magnitude of the chemical shifts of amide- and alpha-protons can be used as a measurement of the complex stability and binding site of the peptide. Our NMR results propose a 3(10)-helical structure for BL-DIS6, and suggest a binding cavity for DPH that involves the hydrophobic particles of residues Ans-7, Leu-8, Val-11, and Val-12. Furthermore, molecular modeling was performed to provide a possible complex conformation that the phenyl portion of DPH is accommodated in the proximity of the C-terminal residues Ala-11 and Val-12, and simultaneously the heterocyclic amine ring of DPH is perching at the residue Asn-7 periphery and stabilizing the phenyl portion deep insertion into the peptide.

The conformation and movement of Na channel inactivation gate peptide in linker between domain III and IV during inactivation by NMR spectroscopy and molecular modeling study.

Amodel peptide that their sequence corresponds to the linker part between domain III and IV of rat brain type IIA Na+ channel has been synthesized for the conformational affect study corresponded to different gated states of Na+ channel. Nuclear magnetic resonance spectra of local anesthetic (LA) diphenyl drugs, such as phenytoin, in presence of a model peptide in both phosphate buffer and phospholipid bicelles (dimyristotl phosphocholine/dihexanoyl phospholcholine), which micelles serve to mimic the peptide-lipid interactions, have been measured to obtain information of the interactions between selected drugs and model peptide. Molecular modeling is performed to help to provide possible conformational information about the polypeptide LIII-IV that may be critical for recognition and signal transduction of inactivated Na+ channel. The voltage-sensing mechanism of Na+ channel involves the movement of the inactivation particles (Ile, Phe, and Met) in the LIII-IV while binding to S4-S5 intracellular region within DIII and DIV. The movement of LIII-IV making its C-terminal residues, including Glu1492 and Glu1493, may aligned near and stabilize the LAs bound with their receptors.

3: Community-acquired pneumonia.

Community-acquired pneumonia is caused by a range of organisms, most commonly Streptococcus pneumoniae, Mycoplasma pneumoniae, Chlamydia pneumoniae and respiratory viruses. Chest x-ray is required for diagnosis. A risk score based on patient age, coexisting illness, physical signs and results of investigations can aid management decisions. Patients at low risk can usually be managed with oral antibiotics at home, while those at higher risk should be further assessed, and may need admission to hospital and intravenous therapy. For S. pneumoniae infection, amoxycillin is the recommended oral drug, while benzylpenicillin is recommended for intravenous use; all patients should also receive a tetracycline (eg, doxycycline) or macrolide (eg, roxithromycin) as part of initial therapy. Flucloxacillin or dicloxacillin should be added if staphylococcal pneumonia is suspected, and gentamicin or other specific therapy if gram-negative pneumonia is suspected; a third-generation cephalosporin plus intravenous erythromycin is recommended as initial therapy for severe cases. Infections that require special therapy should be considered (eg, tuberculosis, melioidosis, Legionella, Acinetobacter baumanii and Pneumocystis carinii infection).

[Introduction of the chromogenic gene to the plant growth-promoting rhizobacteria of cucumber].

Using a bicomponent transposition system with the E. coli lacZY gene cloned between Tn7 termini, a sensitive, selectable marker based on expression of the E. coli lac operon genes encoding beta-galactosidase and lactose permease was transformed into the rifampicin resistant mutant of plant growth-promoting rhizobacteria of cucumber, Pseudomonas aeruginosa CN116 and Pseudomonas corrugata CN31, respectively. Transformants were conferred the ability to utilize lactose as a sole carbon source and the ability to cleave the chromogenis substrate X-Gal to show a specific blue color. Southern blotting analysis showed that lacZY gene was inserted into the genome DNA of target strains. Compared with the wild type strains, the cultural characters, morphological features, growth promoting and disease control effects of transformants were almost unchanged, except the new marked phenotype. This marker system enabled the detection of lac+ transformants at sensitivity of 10 CFU/g soil, which makes the further studies on PGPR more easily.

Resonance Raman studies indicate a unique heme active site in prostaglandin H synthase.

Prostaglandin H synthase isoforms 1 and 2 (PGHS-1 and -2) catalyze the first two steps in the biosynthesis of prostaglandins. Resonance Raman spectroscopy was used to characterize the PGHS heme active site and its immediate environment. Ferric PGHS-1 has a predominant six-coordinate high-spin heme at room temperature, with water as the sixth ligand. The proximal histidine ligand (or the distal water ligand) of this hexacoordinate high-spin heme species was reversibly photolabile, leading to a pentacoordinate high-spin ferric heme iron. Ferrous PGHS-1 has a single species of five-coordinate high-spin heme, as evident from nu(2) at 1558 cm(-1) and nu(3) at 1471 cm(-1). nu(4) at 1359 cm(-1) indicates that histidine is the proximal ligand. A weak band at 226-228 cm(-1) was tentatively assigned as the Fe-His stretching vibration. Cyanoferric PGHS-1 exhibited a nu(Fe)(-)(CN) line at 446 cm(-1) and delta(Fe)(-)(C)(-)(N) at 410 cm(-1), indicating a "linear" Fe-C-N binding conformation with the proximal histidine. This linkage agrees well with the open distal heme pocket in PGHS-1. The ferrous PGHS-1 CO complex exhibited three important marker lines: nu(Fe)(-)(CO) (531 cm(-1)), delta(Fe)(-)(C)(-)(O) (567 cm(-1)), and nu(C)(-)(O) (1954 cm(-1)). No hydrogen bonding was detected for the heme-bound CO in PGHS-1. These frequencies markedly deviated from the nu(Fe)(-)(CO)/nu(C)(-)(O) correlation curve for heme proteins and porphyrins with a proximal histidine or imidazolate, suggesting an extremely weak bond between the heme iron and the proximal histidine in PGHS-1. At alkaline pH, PGHS-1 is converted to a second CO binding conformation (nu(Fe)(-)(CO): 496 cm(-1)) where disruption of the hydrogen bonding interactions to the proximal histidine may occur.

Use of inhibitors to evaluate coreceptor usage by simian and simian/human immunodeficiency viruses and human immunodeficiency virus type 2 in primary cells.

We have used coreceptor-targeted inhibitors to investigate which coreceptors are used by human immunodeficiency virus type 1 (HIV-1), simian immunodeficiency viruses (SIV), and human immunodeficiency virus type 2 (HIV-2) to enter peripheral blood mononuclear cells (PBMC). The inhibitors are TAK-779, which is specific for CCR5 and CCR2, aminooxypentane-RANTES, which blocks entry via CCR5 and CCR3, and AMD3100, which targets CXCR4. We found that for all the HIV-1 isolates and all but one of the HIV-2 isolates tested, the only relevant coreceptors were CCR5 and CXCR4. However, one HIV-2 isolate replicated in human PBMC even in the presence of TAK-779 and AMD3100, suggesting that it might use an undefined, alternative coreceptor that is expressed in the cells of some individuals. SIV(mac)239 and SIV(mac)251 (from macaques) were also able to use an alternative coreceptor to enter PBMC from some, but not all, human and macaque donors. The replication in human PBMC of SIV(rcm) (from a red-capped mangabey), a virus which uses CCR2 but not CCR5 for entry, was blocked by TAK-779, suggesting that CCR2 is indeed the paramount coreceptor for this virus in primary cells.

Inhibition of Na(+) current by diphenhydramine and other diphenyl compounds: molecular determinants of selective binding to the inactivated channels.

Diphenhydramine is an H1 histamine receptor antagonist, yet it also has a clinically useful local anesthetic effect. We found that diphenhydramine inhibits the neuronal Na(+) current, and the inhibition is stronger with more positive holding potentials. The dissociation constant between diphenhydramine and the inactivated Na(+) channel is approximately 10 microM, whereas the dissociation constant between diphenhydramine and the resting channel is more than 300 microM. The local anesthetic effect of diphenhydramine thus is ascribable to inhibition of Na(+) current by selective binding of the drug to the inactivated channels. Most interestingly, many other compounds, such as the anti-inflammatory drug diclofenac, the anticonvulsant drug phenytoin, the antidepressant drug imipramine, and the anticholinergic drug benztropine, have similar effects on neuronal Na(+) current. There is no apparent common motif in the chemical structure of these compounds, except that they all contain two phenyl groups. Molecular modeling further shows that the two benzene rings in all these drugs have very similar spatial orientations (stem bond angle, approximately 110 degrees; center-center distance, approximately 5 A). In contrast, the two phenyl groups in phenylbutazone, a drug that has only a slight effect on Na(+) current, are oriented in quite a different way. These findings strongly suggest that the two phenyl groups are the key ligands interacting with the channel. Because the binding counterpart of a benzene ring usually is also a benzene ring, some aromatic side chain groups of the Na(+) channel presumably are realigned during the gating process to make the very different affinity to the aforementioned drugs between the inactivated and the resting channels.