New alleles in the B44 family including B*44022, B*44032, B*4411, B*4420, B*4421, B*4424, and B*8301

Seven new HLA-B locus alleles have been described. B*44022 and B*44032 are silent substitutions altering known alleles. B*4411 carries a unique Bw4-like epitope. B*4420, B*4421, and B*4424 carry new combinations of motifs previously observed in other alleles. B*8301 appears to be the result of the replacement of exon 2 from B*4402 with exon 2 from B*5603.     

Chemically and electrically coupled interneurons mediate respiratory pumping in Aplysia

1. Respiratory pumping in Aplysia consists of synchronous, brief contractions of the mantle organs that can occur spontaneously and also can be triggered by tactile or noxious stimulation. It has been shown previously to be driven in part by a cluster of electrically coupled interneurons, called the L25 cells, located in the left hemiabdominal ganglion. This paper describes a second class of interneurons, the R25 cells, that also plays an important role in the control of respiratory pumping. 2. The R25 cells are a cluster of approximately 14 interneurons located in the right hemiabdominal ganglion, roughly symmetrical to the L25 cells. The R25 cells are electrically coupled to each other and to the L25 cells. Some R25 and L25 cells also produce chemically mediated fast excitation and slow inhibition of other neurons in the R25/L25 network. 3. Three lines of evidence demonstrate that the R25 cells play a role in mediating respiratory pumping: 1) They fire in a synchronous burst each time the behavior occurs. 2) They make direct chemical synaptic connections to motoneurons that mediate the behavior; and 3) Firing a single R25 cell can trigger the entire behavior, by recruiting synchronous bursts of activity in all of the other R25 and L25 neurons. Individual R25 and L25 cells can act both as trigger cells (exciting the other interneurons) and as relay cells (projecting directly to motoneurons). 4. Burst initiation within the R25/L25 network appears to have two phases: 1) There is an initial phase when the R25 and L25 cells fire at a relatively low frequency. This phase can be driven either by endogenous pacemaker activity of the R25/L25 cells or by afferent synaptic input from sensory pathways; and 2) The late, high-frequency phase of the burst results largely from reverberation within the network, as activity in each cell contributes positive feedback via the excitatory chemical and electrical connections between R25 and L25 cells. 5. Synchronization of the different motor outputs that make up the pumping behavior is achieved by three mechanisms: 1) When pumping occurs spontaneously, the electrical coupling between the cells of the R25/L25 network ensures that these cells will all be near spike threshold at the end of each interburst period.(ABSTRACT TRUNCATED AT 400 WORDS)     

Detection of Chlamydia trachomatis inclusions in Mccoy cell cultures with fluorescein-conjugated monoclonal antibodies.

We compared two methods for identification of Chlamydia trachomatis inclusions in McCoy cell monolayers: conventional iodine staining and immunofluorescence staining with monoclonal antibodies against the species-specific major outer membrane protein antigen of C. trachomatis. Among 878 urethral and cervical specimens tested in parallel, the immunofluorescence method detected eightfold more inclusions per monolayer, identified a higher proportion of positive specimens on first passage (98 versus 62% by iodine staining; P less than 0.01), and improved overall sensitivity (98% of total positive specimens detected versus 84% by iodine staining; P less than 0.01). Improved sensitivity was most evident in specimens with low numbers of inclusions. Compared with conventional iodine staining, immunofluorescence staining with monoclonal antibodies improves sensitivity and offers more rapid detection of chlamydial inclusions in cell culture.     

Twelve novel HLA-B*15 alleles carrying previously observed sequence motifs are placed into B*15 subgroups

Twelve new B*15 alleles are described. All of the known B*15 alleles are divided into subgroups based on serologic assignments and/or nucleotide sequence polymorphisms. These groups might be used as a reference for DNA-based testing at an intermediate (i.e. "serologic") level of resolution.     

Schedule dependent potentiation of antitumor drug effects by α-difluoromethylornithine in human gastric carcinoma cells in vitro

A clone of human gastric cancer cells (AGS-6) and the parental line (AGS-P) from which it was isolated were used in cell survival studies to determine whether pretreatment for 24, 48 or 72h with -difluoromethylornithine (DFMO, 5mM) would increase the cell's sensitivity to 5-Fluorouracil (5FU), Adriamycin (Adria), 1-(2-chloroethyl)-3-(4-methyl cyclohexyl)-1-nitrosourea (MeCCNU), or Bleomycin (Bleo). Generally, the AGS parental cells were most sensitive to the anticancer agents after exposures to DFMO. However, there was no way to predict in advance from DFMO-induced changes in ornithine decarboxylase (ODC), polyamine or cell kinetics values, how long an exposure to DFMO was required before sensitization to an anticancer agent occurred. The degree of potentiation for a single drug was variable from time to time during exposure to DFMO, and broad differences in the sensitizations were demonstrated among the four anticancer drugs. The AGS-6 clone exhibited little or no increased sensitivity as a result of pretreatment with DFMO, even though the DFMO-induced reductions in ODC and polyamine values in these cells were similar to those produced in the more sensitive parental line.     

Flow cytometry: Potential utility in monitoring drug effects in breast cancer

Summary Flow cytometric analysis of DNA ploidy and S-phase fraction are well recognized prognostic indicators in breast cancer. The present paper deals with the widening of the applications of flow cytometry to monitoring the effectiveness of antiestrogen therapy, detecting clonal selection and emergence of drug resistance, and monitoring chemosensitizing properties of drugs. Antiestrogen activity can be studied by DNA flow cytometry to address clinical research problems such as patient-specific pharmacokinetics, dosing compliance, and acquired antiestrogen resistance. Patient plasma specimens containing various concentrations of triphenylethylenes can be monitored for drug-induced effects using cell cycle measurements and correlated toin vivo drug levels. DNA flow cytometry has also been instrumental in the study of the effects of prolonged low-dose (0.5 µM for > 100 days) tamoxifen treatment on human estrogen receptor negative MDA-MB-231 cells, where it was shown that tamoxifen may significantly alter cell cycle kinetics and tumorigenicity of these cells, selecting a new, more aggressive, and rapidly growing clone. Lastly, it has been shown that the chemosensitizing properties of another triphenylethylene antiestrogen, toremifene, on estrogen receptor negative, multidrug resistant MDA-MB-231-A1 human breast cancer cells can be studied using flow cytometric analysis. Toremifene (and its metabolites N-desmethyltoremifene and toremifene IV) are able to resensitize MDA-MB-231-A1 cells to vinblastine and doxorubicin, as reflected in a marked shift of cells to G₂/M phase of the cell cycle. Flow cytometry is a widely available technique that might be applied clinically to monitor, at the cellular level, drug effects on tumors, including the modulators of drug resistance.     

Intravenous drug users and AIDS: risk behaviors

Risk-taking behaviors were studied in this assessment of 345 intravenous drug users from Baltimore, El Paso, and Denver. Over 50% reported injecting drugs daily and 70% shared needles with others, averaging 6.3 partners. In addition, 86% had shared a "cooker" and nearly 50% injected in a "shooting gallery." More than half of the males sampled had two or more sex partners, including 18% with five or more. Females averaged 19 sex partners in the preceding 6 months, with 22% reporting sex with five or more. Two-thirds of the total sample never used a condom, while only 6% always used this form of protection. On the other end of this risk continuum were those subjects who did not share needles or always cleaned their needles with an effective agent, had no sexual relations or always used a condom. Subjects following such practices could be considered low risk if they adopted safe behaviors in other associated areas of their lives. However, in an analysis of total risk, it was found that only 14 subjects (4%) practiced safe needle use and safe sex. Despite these findings, some encouraging results were seen. In an analysis of risk according to location, Baltimore subjects were significantly less at risk according to number of needle-sharing partners, borrowing needles, sharing a "cooker," injection in a "shooting gallery," cleaning needles, use of disinfectants, number of sexual partners, and use of condoms than either their cohorts in El Paso or Denver. Street outreach to modity risk behaviors among IVDUs began in Baltimore approximately 2 years prior to funding in El Paso and Denver. These results suggest that there may be a potential to moderate risk through intervention.     

Head and neck tumors: imaging recurrent tumor and post-therapeutic changes with CT and MRI

OBJECTIVE: To evaluate criteria for detection of tumor recurrence and post-treatment changes in patients with head and neck malignancies in computed tomography (CT) and magnetic resonance imaging (MRI). METHODS AND MATERIALS: Thirty-nine patients with head and neck carcinoma receiving radiochemotherapy were examined before, during and after therapy with MRI. Changes in signal intensity were correlated to histology or clinical course. Three hundred and thirty-one patients with head and neck malignancies were examined with CT after therapy. CT diagnoses were correlated with histology or clinical course. RESULTS: Main criteria for recurrent/residual tumor in MRI was infiltrative mass with high signal intensity in T2-weighted images and enhancement after Gd-DTPA in T1-weighted images. Radiation-induced changes led to false positive diagnosis in 46% in the interval up to 3 months after therapy and in 58% in the interval 3-6 months after therapy. The combination of a circumscribed, infiltrative mass with contrast enhancement in CT had a sensitivity of 86% and a specificity of 80%. CONCLUSION: CT could accurately demonstrate postoperative changes and tumor recurrence. MRI had advantages in differentiation of tumor and scar, but edema after radiation therapy can spoil diagnosis.     

Heroin and Methamphetamine Injection: An Emerging Drug Use Pattern

Objective: We sought to describe an emerging drug use pattern characterized by injection of both methamphetamine and heroin. We examined differences in drug injection patterns by demographics, injection behaviors, HIV and HCV status, and overdose. Methods: Persons who inject drugs (PWID) were recruited as part of the National HIV Behavioral Surveillance (NHBS) system in Denver, Colorado. We used chi-square statistics to assess differences between those who reported only heroin injection, only methamphetamine injection, and combined heroin and methamphetamine injection. We used generalized linear models to estimate unadjusted and adjusted prevalence ratios to describe the association between drug injection pattern and reported nonfatal overdose in 2015. We also examined changes in the drug reported as most frequently injected across previous NHBS cycles from 2005, 2009, and 2012. Results: Of 592 participants who completed the survey in 2015, 173 (29.2%) reported only injecting heroin, 123 (20.8%) reported only injecting methamphetamine, and 296 (50.0%) reported injecting both drugs during the past 12 months. Injecting both heroin and methamphetamine was associated with a 2.8 (95% confidence interval: 1.7, 4.5) fold increase in reported overdose in the past 12 months compared with only injecting heroin. The proportion of those reporting methamphetamine as the most frequently injected drug increased from 2.1% in 2005 to 29.6% in 2015 (p < 0.001). Conclusions: The rapid increase in methamphetamine injection, and the emergence of combining methamphetamine with heroin, may have serious public health implications.     

Intrasomatic injection of radioactive precursors for studying transmitter synthesis in identified neurons of Aplysia californica

We introduced radioactive precursors directly into identified neurons of Aplysia californica. [(3)H]-Choline and L-[(3)H]tryptophan were injected with pressure into nerve cell bodies to study synthesis of acetylcholine and serotonin. We confirmed the cholinergic nature of R2, L10, and L11, identified neurons of the abdominal ganglion. Cells in the LD cluster (which contains motor neurons to the heart and gill) also converted most of the injected choline into acetylcholine. Neurons in the RB cluster (which contains an excitatory motor neuron to the heart) and the two metacerebral cells of the cerebral ganglion converted injected tryptophan to serotonin. No cell studied could convert both choline to acetylcholine and tryptophan to serotonin. Pressure permits rapid injection of precursors, from small amounts to amounts large enough to saturate intracellular synthetic pathways. In contrast to the results with injection, we found far less synthesis of acetylcholine and serotonin in identified nerve cell bodies when ganglia were incubated in the presence of the radioactive precursors.