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Objective: To evaluate whether less pulsatile levodopa therapy (LPT) can reduce the development of levodopa-induced dyskinesia (LID)., Methods: This is a retrospective cohort study of patients with Parkinson's disease at the movement disorders clinic of Medstar Washington Hospital Center. The study was not blinded or randomized. Patients were seen between August 2002 and August 2018. During these years, we treated patients with less pulsatile (6 doses daily) levodopa treatment to reduce LID. Occurrence of LID was recorded., Results: Ninety-five patients with Parkinson's disease taking levodopa were divided into two groups: 1) patients who were initially managed on LPT or who switched from traditional therapy (TT) (n = 61) (mean disease duration: 7.7 ± 4.8 years, mean levodopa duration: 5.6 ± 4.5 years and mean observation time: 4.3 ± 3.4 years), and 2) patients on TT throughout the observation period or until they developed dyskinesia (n = 34) (mean disease duration: 8.3 ± 3.8 years, mean levodopa duration: 6.2 ± 4.2 years and mean observation time: 4.1 ± 3.4 years). Three of the 61 LPT patients developed dyskinesia during the observation period. One of the patients developed dyskinesia after being switched to pulsatile doses by another doctor. In the other two, dyskinesia was minimal. In contrast to this 4.9% cumulative incidence, dyskinesia occurred in 50% (17/34) of TT patients, an incidence similar to that in published data (p < 0.001)., Conclusion: Less pulsatile levodopa with 6 daily doses was associated with a low incidence of LID. Further study of this method of treatment is warranted.

Individualized medicine is a new direction in the field of modern pharmacy. In this study, we assessed the feasibility and accuracy of 3D printing techniques for the preparation of individualized doses of mouth-disintegrating tablets of warfarin. Warfarin sodium, D-sucrose, pregelatinized starch, povidone K30, microcrystalline cellulose, and silicon dioxide (at a ratio of 1:42.45:46.15:5.1:4.9:0.4) were mixed and used as the printing powder in the 3D printer; preset parameters were used. The dosage of the tablet was controlled by the number of printing layers. The content, dose uniformity, dose accuracy, hardness, friability, disintegration time, dissolution, and the microstructural and overall appearance were determined to evaluate the printed tablets. For the doses of 3, 2, and 1 mg that were produced in the experiment, the disintegration times were 50.0 ± 5.2, 35.7 ± 4.3, and 11.0 ± 2.2 s, respectively, and the relative errors of the dose were -2.33, -1.50, and 0%, respectively. The other indicators were consistent with the preparation requirements of pharmaceutical tablets. It is possible to prepare tablets with excellent properties and controlled drug doses by using 3D printing techniques. This technology will be an important means to achieve individualized medicine.

Purpose: We evaluate diabetic microaneurysm (MA) features on high-resolution adaptive optics scanning laser ophthalmoscopy (AOSLO) and their correlations with visual acuity (VA) and local retinal pathology on spectral domain optical coherence tomography (SDOCT)., Methods: Diabetic participants underwent VA testing and AOSLO and SDOCT imaging of MAs. AOSLO images were graded for MA dimension, wall hyperreflectivity (WH), intraluminal hyperreflectivity (IH), and perfusion pattern. SDOCTs centered on each MA were graded for disorganization of the retinal inner layers (DRIL) and other neuroretinal pathology., Results: We imaged 109 MAs (30 eyes). Multivariate modeling, including statistically significant covariates from bivariate analyses, associated WH with greater MA size (P = 0.001) and DRIL (P = 0.04). IH was associated with perfusion (P = 0.003) and MA visibility on photographs (P = 0.0001), and larger MA size with partial perfusion (P = 0.03), MA ring signs (P = 0.0002), and photographic visibility (P = 0.01). Multivariate modeling revealed an association of WH and VA with DRIL., Conclusions: AOSLO imaging demonstrates associations of hyperreflective MA walls with MA size and adjacent DRIL, as well as the presence of DRIL with lower VA. This study identifies a correlation between vascular and neural pathology associated with VA decline. Further studies of MA structure and neuroretinal disorganization may enable novel approaches to assess anatomic and functional outcomes in the diabetic eye.

Understanding the attitudes and willingness in regard to living organ donation of health care professionals is important for clinical practice and needs to be addressed. Thus, the objective of this study was to examine willingness and its influencing factors in regard to living organ donation in clinical health care professionals., Methods: This study used a cross-sectional design. A total of 375 health care professionals from a regional teaching hospital in northern Taiwan were included in this study., Results: Nearly 65% of the participants indicated a willingness to donate their living organ. Of these participants, 97.1% were willing to donate to family members, 80.8% were willing to donate to friends, and 78.9% were willing to donate to strangers. The predictors of willingness to engage in living organ donation were the desire to help others (odds ratio [OR] = 2.96; P < .01), positive attitude toward living organ donation (OR = 1.12; P < .01), financial support from the government (OR = 4.99; P < .01), and fewer physical concerns (OR = 0.97; P = .04). The willingness to donate a living organ was not associated with age, sex, religious belief, education level, participation in voluntary work, years of clinical work, type of profession, or knowledge about living organ donation., Conclusion: In general, health care professionals had a positive attitude toward and willingness to engage in living organ donation. It is hoped that the results of this study will serve as a referent framework for policymaking in regard to living organ donation and transplantation., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Purpose: Understanding patients' attitudes toward novel therapeutic options can help guide providers in personalizing treatment regimens for glaucoma patients. This study aimed to determine factors associated with acceptance of new drug delivery options among glaucoma patients., Methods: A total of 199 patient volunteers participated in an interviewer-administered survey from June to August 2016 at the Glaucoma Service of Massachusetts Eye and Ear. The questionnaire was designed to determine acceptance of 6 drug delivery approaches: (1) triple combination eye drop, (2) microdose eye spray, (3) drug-eluting contact lens, (4) drug-eluting periocular ring insert, (5) injectable subconjunctival drug insert, and (6) injectable anterior chamber implant. Other factors analyzed included self-reported demographics, disease severity, and prior ocular history., Results: The average respondent age was 63.2 ± 15.1 years; 48% were female. For approaches 1-6 listed above, overall acceptance rates were, respectively, 85%, 54%, 31%, 43%, 32%, and 30%. Patients with greater disease severity and prior incisional glaucoma surgery were more likely to pursue alternatives to traditional eye drops., Conclusions: There is limited acceptance of alternatives to traditional eye drop medications among glaucoma patients. Understanding motivating factors and potential barriers to patient acceptance of novel drug delivery approaches is important in how providers will incorporate these glaucoma treatment options into practice.

This study describes the synthesis of novel gene delivery vector with low toxicity and high transfection efficiency for magnetofection. The rational design of magnetofection vector called PPMag (PEI-associated polycaprolactone (PCL)-SPIONs) composed of oleic acid (OA) stabilized superparamagnetic iron oxide nanoparticles (SPPIONs) prepared by thermolysis of iron oleate with a combination of hydrophobic PCL and proton absorbing polymer polyethyleneimine (PEI) (PEI-PCL-SPIONs) is described. Encapsulation of amphiphilic PEI with SPIONs not only improves water dispersity of SPIONs, but also allows nucleic acid (NA) condensation and endosomal/lysosomal escape via proton sponge effect after internalization in cells. MTT cytotoxicity assay showed that cell viability was improved compared to conventional PEI-SPIONs. The luciferase activity of magneto-polyplexes treated cells significantly improved compared to both controls revealed that transfection efficiency of PPMag- pCIKlux polyplexes group was improved compared to naked pCIKlux group. The application underneath of a rare earth magnet significantly improve the transfection efficiency (i.e., the luciferase activity doubles) compared to cells without magnet, indicating that sedimentation induced by magnetic field plays important role in accumulation of magneto-polyplexes on cell surfaces. The results demonstrate that PPMag can be used as a novel gene transfection vector to improve transfection efficiency. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 145-154, 2017., (© 2015 Wiley Periodicals, Inc.)

In this study, we investigated the effects of a symbiotic bacterium and phosphate (PO4(3-)) nutrition on the toxicity and metabolism of arsenate (As(V)) in Dunaliella salina. The bacterium was identified as Alteromonas macleodii based on analysis of its 16S rRNA gene sequence. When no As(V) was added, A. macleodii significantly enhanced the growth of D. salina, irrespective of PO4(3-) nutrition levels, but this effect was reversed after As(V)+PO4(3-) treatment (1.12mgL(-1)) for 3 days. Arsenic (As) absorption by the non-axenic D. salina was significantly higher than that by its axenic counterpart during incubation with 1.12mgL(-1) PO4(3-). However, when the culture was treated with 0.112mgL(-1) PO4(3-), As(V) reduction and its subsequent arsenite (As(III)) excretion by non-axenic D. salina were remarkably enhanced, which, in turn, contributed to lower As absorption in non-axenic algal cells from days 7 to 9. Moreover, dimethylarsinic acid was synthesized by D. salina alone, and the rates of its production and excretion were accelerated when the PO4(3-) concentration was 0.112mgL(-1). Our data demonstrate that A. macleodii strongly affected As toxicity, uptake, and speciation in D. salina, and these impacts were mediated by PO4(3-) in the cultures., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The present report proposes a more rational hyaluronic acid (HA) conjugation protocol that can be used to modify the surface of the superparamagnetic iron oxide nanoparticles (SPIONs) by covalently binding the targeting molecules (HA) with glutamic acid as a molecular linker on peripheral surface of SPIONs. The synthesis of HA-Glutamic Acid (GA)@SPIONs was included oxidization of nanoparticle's surface with H₂O₂ followed by activation of hydroxyl group and reacting glutamic acid as an intermediate molecule demonstrating transfection of lung cancer cells. Fourier transform infrared (FTIR) and zeta-potential studies confirmed the chemical bonding between amino acid linker and polysaccharides. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cytotoxicity assay showed that HA-SPIONs-treated cells remained 82.9% ± 2.7% alive at high particle dosage (200 µg/mL iron concentration), whereas GA-SPIONs and bare SPIONs (B-SPIONs) treated cells had only 59.3% ± 13.4% and 26.5% ± 3.1% survival rate at the same conditions, respectively. Confocal microscopy analysis showed increased cellular internalization of HA-SPIONs compared to non-interacting agarose coated SPIONs (AgA-SPIONs).

A prospective cohort study was conducted to determine whether chromosome aneuploidy increases the risk of early spontaneous abortions in patients with polycystic ovary syndrome (PCOS). A total of 1461 patients who conceived after IVF and embryo transfer were followed; 100 patients who had experienced clinical spontaneous abortion were recruited, 32 with PCOS and 68 without PCOS. Before 2013, genetic analysis comprised conventional cultured villus chromosome karyotyping and a multiplex ligation-dependent probe amplification subtelomere assay combined with fluorescence in-situ hybridization; since 2013, array-based comparative genomic hybridization technique combined with chromosome karyotyping has been used. Age, BMI, pregnancy history, gestational age and total gonadotrophin dosage did not differ significantly between the PCOS and non-PCOS groups. In the PCOS group, 28.1% of abortuses demonstrated aneuploidy, which was significantly lower (P = 0.001) than in the non-PCOS group (72.1%). Further statistical analyses controlling for maternal age demonstrated that abortuses of women with PCOS were significantly less (P = 0.001) likely to have chromosome aneuploidy. Embryonic aneuploidy does not play a vital role in early spontaneous abortion in women with PCOS. Maternal factors resulting in endometrial disorders are more likely to be responsible for the increased risk of early spontaneous abortion in patients with PCOS., (Copyright © 2016 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.)

We describe a patient who underwent coronary artery bypass grafting (CABG) surgery with the presentation of acute coronary syndrome (ACS). The diagnostic coronary angiogram showed acute thrombotic and occluded saphenous vein graft (SVG) and proximal right coronary artery (RCA) drug eluting stent (DES) instent restenosis (ISR) with chronic total occlusion (CTO). Our strategy was to recanalize the native left circumflex coronary artery (LCx) CTO instead of SVG or RCA instent CTO. After heparinization for 5 days, the LCx antegrade approach and the retrograde approach from left anterior descending coronary artery (LAD) septal branches were first attempted but failed, and the LCx CTO was successfully revascularized retrogradely via the acute thrombotic SVG without an embolic protection device (EPD).

KAE609 [(1'R,3'S)-5,7'-dichloro-6'-fluoro-3'-methyl-2',3',4',9'-tetrahydrospiro[indoline-3,1'-pyridol[3,4-b]indol]-2-one] is a potent, fast-acting, schizonticidal agent being developed for the treatment of malaria. After oral dosing of KAE609 to rats and dogs, the major radioactive component in plasma was KAE609. An oxidative metabolite, M18, was the prominent metabolite in rat and dog plasma. KAE609 was well absorbed and extensively metabolized such that low levels of parent compound (≤11% of the dose) were detected in feces. The elimination of KAE609 and metabolites was primarily mediated via biliary pathways (≥93% of the dose) in the feces of rats and dogs. M37 and M23 were the major metabolites in rat and dog feces, respectively. Among the prominent metabolites of KAE609, the isobaric chemical species, M37, was observed, suggesting the involvement of an isomerization or rearrangement during biotransformation. Subsequent structural elucidation of M37 revealed that KAE609, a spiroindolone, undergoes an unusual C-C bond cleavage, followed by a 1,2-acyl shift to form a ring expansion metabolite M37. The in vitro metabolism of KAE609 in hepatocytes was investigated to understand this novel biotransformation. The metabolism of KAE609 was qualitatively similar across the species studied; thus, further investigation was conducted using human recombinant cytochrome P450 enzymes. The ring expansion reaction was found to be primarily catalyzed by cytochrome P450 (CYP) 3A4 yielding M37. M37 was subsequently oxidized to M18 by CYP3A4 and hydroxylated to M23 primarily by CYP1A2. Interestingly, M37 was colorless, whereas M18 and M23 showed orange yellow color. The source of the color of M18 and M23 was attributed to their extended conjugated system of double bonds in the structures., (Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.)

How protein structure encodes functionality is not fully understood. For example, long-range intraprotein communication can occur without measurable conformational change and is often not captured by existing structural correlation functions. It is shown here that important functional information is encoded in the timing of protein motions, rather than motion itself. I introduce the conditional activity function to quantify such timing correlations among the degrees of freedom within proteins. For three proteins, the conditional activities between side-chain dihedral angles were computed using the output of microseconds-long atomistic simulations. The new approach demonstrates that a sparse fraction of side-chain pairs are dynamically correlated over long distances (spanning protein lengths up to 7 nm), in sharp contrast to structural correlations, which are short-ranged (<1 nm). Regions of high self- and inter-side-chain dynamical correlations are found, corresponding to experimentally determined functional modules and allosteric connections, respectively.

Hedgehog (Hh) pathway activation in R26-SmoM2;CAGGS-CreER mice, which carry a tamoxifen-inducible activated Smoothened allele (SmoM2), results in numerous microscopic tumor foci in mouse skeletal muscle. These tumors exhibit a highly differentiated myogenic phenotype and resemble human fetal rhabdomyomas. This study sought to apply previously established strategies to isolate lineally distinct populations of normal mouse myofiber-associated cells in order to examine cellular heterogeneity in SmoM2 tumors. We demonstrate that established SmoM2 tumors are composed of cells expressing myogenic, adipocytic and hematopoietic lineage markers and differentiation capacity. SmoM2 tumors thus recapitulate the phenotypic and functional hetereogeneity observed in normal mouse skeletal muscle. SmoM2 tumors also contain an expanded population of PAX7+ and MyoD+ satellite-like cells with extremely low clonogenic activity. Selective activation of Hh signaling in freshly isolated muscle satellite cells enhanced terminal myogenic differentiation without stimulating proliferation. Our findings support the conclusion that SmoM2 tumors represent an aberrant skeletal muscle state and demonstrate that, similar to normal muscle, myogenic tumors contain functionally distinct cell subsets, including cells lacking myogenic differentiation potential., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Rich1, a previously identified Rho GTPase-activating protein (RhoGAP), was found to have close relationship with Rho GTPase family members in multiple cellular processes in nervous cells and platelets. But the exact role of Rich1 in epithelial cells remains obscure. The present investigation demonstrated that up-regulation of Rich1 could cause S-phase arrest, proliferation inhibition and adhesion decline with F-actin amount decrease in epithelial cells. Further exploration in hepatocyte HL7702 revealed that overexpression of Rich1 could greatly elevate the intrinsic GTPase activities on both of CDC42 and RAC1 by stimulating GTP hydrolysis, which consequently attenuated the activities of the Rho proteins and the phosphorylation level of those in PAK1-ERK1/2 signaling cascade. While the GAP domain deleted Rich1 variant or silence of endogenous Rich1 expression could not result in any of the biological effects. It is indicated that Rich1, completely different from in other types of cells, might act as a crucial upstream negative regulator via its GAP domain in control of epithelial cell cycle, proliferation and focal adhesion through CDC42/RAC1-PAK1-ERK1/2 signaling pathway and F-actin dynamics., (Copyright © 2015. Published by Elsevier Inc.)

Despite treatment advances, diabetic eye disease remains a leading cause of visual acuity (VA) loss worldwide. No methods to prospectively determine which patients will gain or lose vision exist, limiting individualized risk assessment and management. We investigated whether noninvasive, readily obtainable spectral domain optical coherence tomography parameters were correlated with VA in eyes with current or resolved center-involved diabetic macular edema (DME). Images were evaluated for disorganization of the retinal inner layers (DRIL), cysts, epiretinal membranes, microaneurysms, subretinal fluid, and outer layer disruption/reflectivity. DRIL affecting ≥50% of the 1-mm central retinal zone was associated with worse VA in all eyes, eyes with current edema, and eyes with resolved edema. Furthermore, early 4-month change in DRIL extent predicted VA change from baseline to 1 year. These data suggest that DRIL is a robust predictor of VA in eyes with present or previous DME and more highly correlated with VA than other widely used measures, such as retinal thickness. If further studies confirm DRIL as a predictive biomarker of future VA, physicians would gain a new tool of substantial clinical and investigative importance that could significantly change the approach to ophthalmic counseling and therapeutic management in patients with diabetes., (© 2015 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.)

Cooperative interactions are widespread in biochemical networks, providing the nonlinear response that underlies behavior such as ultrasensitivity and robust switching. We introduce a temporal correlation function-the conditional activity-to study the behavior of these phenomena. Applying it to the bistable genetic switch in bacteriophage lambda, we find that cooperative binding between binding sites on the prophage DNA lead to non-Markovian behavior, as quantified by the conditional activity. Previously, the conditional activity has been used to predict allosteric pathways in proteins; here, we show that it identifies the rare unbinding events which underlie induction from lysogeny to lysis.

Purpose: Up to 25% of chronic hepatitis B (CHB) patients eventually develop hepatocellular carcinoma (HCC), a disease with poor prognosis unless detected early. This study identifies a blood-based RNA biomarker panel for early HCC detection in CHB., Materials and Methods: A genome-wide RNA expression study was performed using RNA extracted from blood samples from Malaysian patients (matched HCC, CHB, controls). Genes differentiating HCC from controls were selected for further testing using quantitative real-time polymerase chain reaction. Finally, a 6-gene biomarker panel was identified and characterized using a training set (cohort I = 126), and tested against 2 test sets (cohort II = 222; cohort III = 174). The total number of samples used for each group is: HCC + CHB = 143, CHB = 211, control = 168., Results: Our gene panel displays a consistent trend distinguishing HCC from controls in our test sets, with an area under receiver-operating characteristic curve of 0.9 in cohort III. Our independent test set (cohort III) showed that the gene panel had a sensitivity of 70% with a specificity of 92%. The biomarker profile for HCC was consistently detected in a small subgroup of CHB patients, thus potentially predicting early, preclinical cases of cancer that should be screened more intensively., Conclusion: The biomarkers identified in this study can be used as the basis of a blood-based test for the detection of early HCC in CHB.

Importance: Biomarkers that predict future visual acuity (VA) in eyes with baseline diabetic macular edema (DME) would substantively improve risk assessment, management decisions, and selection of eyes for clinical studies targeting DME., Objective: To determine whether baseline or early change in the novel spectral domain-optical coherence tomography (SD-OCT) parameter disorganization of the retinal inner layers (DRIL) is predictive of VA in eyes with center-involved DME., Design, Setting, and Participants: At a tertiary care referral center for diabetic eye disease, a retrospective, longitudinal cohort study obtained demographics, VA, and SD-OCT images from baseline, 4-month, and 8-month visits in 96 participants (120 eyes) with diabetes mellitus and baseline center-involved DME (SD-OCT central subfield thickness, ≥ 320 µm for men and ≥ 305 µm for women). Exclusion criteria included substantial media opacity, cataract surgery within 6 months, and nondiabetic retinal pathology affecting VA. On SD-OCT, the 1-mm-wide retinal area centered on the fovea was evaluated by masked graders for DRIL extent, cysts, hyperreflective foci, microaneurysms, cone outer segment tip visibility, and external limiting membrane or photoreceptor disruption and reflectivity., Main Outcomes and Measures: Visual acuity and SD-OCT-derived retinal morphology., Results: Greater DRIL extent at baseline correlated with worse baseline VA (point estimate, 0.04; 95% CI, 0.02-0.05 per 100 µm; P < .001). An increase in DRIL during 4 months was associated with VA worsening at 8 months (point estimate, 0.03; 95% CI, 0.02-0.05 per 100 µm; P < .001). A multivariate model that included a 4-month change in VA, DRIL, and external limiting membrane disruption was predictive of an 8-month VA change (r = 0.80). Each approximately 300-µm DRIL increase during 4 months predicted a 1-line, 8-month VA decline. When DRIL increased at least 250 µm at 4 months, no eyes had VA improvement of at least 1 line at 8 months. When DRIL decreased at least 250 µm at 4 months, no eyes had VA decline of at least 1 line at 8 months, and 77.7% had VA improvement of at least 1 line., Conclusions and Relevance: Disorganization of the retinal inner layers in the 1-mm foveal area is associated with VA, and change in DRIL predicts future change in VA. Early change in DRIL prospectively identifies eyes with a high likelihood of subsequent VA improvement or decline. Therefore, DRIL warrants further study as a robust, readily obtained, and noninvasive biomarker of future VA response in eyes with DME.

Helices are the "hydrogen atoms" of biomolecular complexity; the DNA/RNA double hairpin and protein α-helix ubiquitously form the building blocks of life's constituents at the nanometer scale. Nevertheless, the formation processes of these structures, especially the dynamical pathways and rates, remain challenging to predict and control. Here, we present a general analytical method for constructing dynamical free-energy landscapes of helices. Such landscapes contain information about the thermodynamic stabilities of the possible macromolecular conformations, as well as about the dynamic connectivity, thus enabling the visualization and computation of folding pathways and timescales. We elucidate the methodology using the folding of polyalanine, and demonstrate that its α-helix folding kinetics is dominated by misfolded intermediates. At the physiological temperature of T = 298 K and midfolding time t = 250 ns, the fraction of structures in the native-state (α-helical) basin equals 22%, which is in good agreement with time-resolved experiments and massively distributed, ensemble-convergent molecular-dynamics simulations. We discuss the prominent role of β-strand-like intermediates in flight toward the native fold, and in relation to the primary conformational change precipitating aggregation in some neurodegenerative diseases.

In 1949, asterixis was first described in patients with hepatic encephalopathy. It was quickly recognized that this phenomenon also occurs in other generalized encephalopathies and sometimes results from structural brain lesions. This paper is a study of asterixis in the general neurology clinic and on the inpatient neurology consultation service. The neurologists recorded the findings on inpatients and clinic patients for 12 consecutive months. Of the 1,109 inpatients with adequate examination, asterixis was documented in 97. Eighteen of the 97 cases were unilateral (18.6%) and 79 cases were bilateral (81.4%). Of the 614 outpatient visits with well documented examination, 6 (1%) individuals had asterixis. Since a small number of patients were examined more than once, the study yielded 103 individuals with adequate data for analysis. Asterixis resulted from varied causes: medications, renal disorder, hepatic dysfunction, pulmonary insufficiency, stroke and other brain lesions (including malignancy, subdural hematoma, and epidural abscess). Asterixis occurred in various patterns: in some cases it was easier to elicit in the upper extremities, in some it was easier to elicit in the lower limbs, and some it was solely or predominantly unilateral. The findings are discussed in light of the literature on asterixis with regard to its varied causes, patterns and presentations. Lastly, asterixis is examined from a historical perspective and the terminology is elucidated.

Severe acute malnutrition (SAM) is a common condition that kills children and intellectually maims those who survive. Close to 20 million children under the age of 5 years suffer from SAM globally, and about 1 million of them die each year. Much of this burden takes place in Asia. Six countries in Asia together have more than 12 million children suffering from SAM: 0.6 million in Afghanistan, 0.6 million in Bangladesh, 8.0 million in India, 1.2 million in Indonesia, 1.4 million in Pakistan, and 0.6 million in Yemen. This article is based on a review of SAM burden and intervention programs in Asian countries where, despite the huge numbers of children suffering from the condition, the coverage of interventions is either absent on a national scale or poor. Countries in Asia have to recognize SAM as a major problem and mobilize internal resources for its management. Screening of children in the community for SAM and appropriate referral and back referral require good health systems. Improving grassroots services will not only contribute to improving management of SAM, it will also improve infant and young child feeding and nutrition in general. Ready-to-use therapeutic food (RUTF), the key to home management of SAM without complications, is still not endorsed by many countries because of its unavailability in the countries and its cost. It should preferably be produced locally from locally available food ingredients. Countries in Asia that do not have the capacity to produce RUTF from locally available food ingredients can benefit from other countries in the region that can produce it. Health facilities in all high-burden countries should be staffed and equipped to treat children with SAM. A continuous cascade of training of health staff on management of SAM can offset the damage that results from staff attrition or transfers. The basic nutrition interventions, which include breastfeeding, appropriate complementary feeding, micronutrient supplementation, and management of acute malnutrition, should be scaled up in Asian countries that are plagued with the burden of malnutrition.

Objective: To investigate the clinical characteristics of Segond fracture and its operative method and opportunity., Methods: From June 2008 to December 2011, arthroscopic exploration was performed in 16 patients with Segond fracture. Six patients were explored at 1 week after injury ,their anterior cruciate ligament were broken completely combined with meniscus injury. Ten patients were explored at 8-10 weeks after injury, 4 patients with anterior and posterior cruciate ligament breakage completely, 4 patients with anterior cruciate ligament breakage completely,and 2 patients with anterior cruciate ligament breakage incompletely,among 10 cases, 8 cases of anterior cruciate ligament breakage completely combined with meniscus injury, 1 case of anterior cruciate ligament breakage incompletely without obviously meniscus injury, 1 case combined with fibular head fracture and lateral collateral ligament injury. All broken cruciate ligaments were rebuilt after arthroscopic exploration and meniscus injuries were sutured in 5 cases. Clinical effects were evaluated according to Lysholm-Gillquist scoring of knee joint function., Results: Six patients were operated at 1 week after injury,their knee joint swelled obviously at 3 days after operation,unloaded blood oozing by joint puncture,and out-of-bed activity with assistance at the I week after operation. Other 10 patients were operated at 8-10 weeks after injury,no knee joint obviously swelled,no blood oozing was found by joint puncture ,and out-of-bed activity with assistance at 3 days after operation. All patients were followed up from 12 to 50 months with an average of 24 months. Postoperative Lysholm-Gillquist scoring of all patients were higher than preoperative and recovered well., Conclusion: Segond fractures often associated with anterior cruciate ligament breakage and meniscus injury,it is important that early detection for treatment. The best time of cruciate ligament rebuilding and meniscus repairing may be at 8-10 weeks after injury.

Background: Treatment protocols for nasopharyngeal carcinoma (NPC) developed in the past decade have significantly improved patient survival. In most NPC patients, however, the disease is diagnosed at late stages, and for some patients treatment response is less than optimal. This investigation has two aims: to identify a blood-based gene-expression signature that differentiates NPC from other medical conditions and from controls and to identify a biomarker signature that correlates with NPC treatment response., Methods: RNA was isolated from peripheral whole blood samples (2 x 10 ml) collected from NPC patients/controls (EDTA vacutainer). Gene expression patterns from 99 samples (66 NPC; 33 controls) were assessed using the Affymetrix array. We also collected expression data from 447 patients with other cancers (201 patients) and non-cancer conditions (246 patients). Multivariate logistic regression analysis was used to obtain biomarker signatures differentiating NPC samples from controls and other diseases. Differences were also analysed within a subset (n=28) of a pre-intervention case cohort of patients whom we followed post-treatment., Results: A blood-based gene expression signature composed of three genes - LDLRAP1, PHF20, and LUC7L3 - is able to differentiate NPC from various other diseases and from unaffected controls with significant accuracy (area under the receiver operating characteristic curve of over 0.90). By subdividing our NPC cohort according to the degree of patient response to treatment we have been able to identify a blood gene signature that may be able to guide the selection of treatment., Conclusion: We have identified a blood-based gene signature that accurately distinguished NPC patients from controls and from patients with other diseases. The genes in the signature, LDLRAP1, PHF20, and LUC7L3, are known to be involved in carcinoma of the head and neck, tumour-associated antigens, and/or cellular signalling. We have also identified blood-based biomarkers that are (potentially) able to predict those patients who are more likely to respond to treatment for NPC. These findings have significant clinical implications for optimizing NPC therapy.

The resistance of methicillin-resistant Staphylococcus aureus (MRSA) to all β-lactam classes limits treatment options for serious infections involving this organism. Our goal is to discover new agents that restore the activity of β-lactams against MRSA, an approach that has led to the discovery of two classes of natural product antibiotics, a cyclic depsipeptide (krisynomycin) and a lipoglycopeptide (actinocarbasin), which potentiate the activity of imipenem against MRSA strain COL. We report here that these imipenem synergists are inhibitors of the bacterial type I signal peptidase SpsB, a serine protease that is required for the secretion of proteins that are exported through the Sec and Tat systems. A synthetic derivative of actinocarbasin, M131, synergized with imipenem both in vitro and in vivo with potent efficacy. The in vitro activity of M131 extends to clinical isolates of MRSA but not to a methicillin-sensitive strain. Synergy is restricted to β-lactam antibiotics and is not observed with other antibiotic classes. We propose that the SpsB inhibitors synergize with β-lactams by preventing the signal peptidase-mediated secretion of proteins required for β-lactam resistance. Combinations of SpsB inhibitors and β-lactams may expand the utility of these widely prescribed antibiotics to treat MRSA infections, analogous to β-lactamase inhibitors which restored the utility of this antibiotic class for the treatment of resistant Gram-negative infections.

To find the native conformation (fold), proteins sample a subspace that is typically hundreds of orders of magnitude smaller than their full conformational space. Whether such fast folding is intrinsic or the result of natural selection, and what is the longest foldable protein, are open questions. Here, we derive the average conformational degeneracy of a lattice polypeptide chain in water and quantitatively show that the constraints associated with hydrophobic forces are themselves sufficient to reduce the effective conformational space to a size compatible with the folding of proteins up to approximately 200 amino acids long within a biologically reasonable amount of time. This size range is in general agreement with the experimental protein domain length distribution obtained from approximately 1,200 proteins. Molecular dynamics simulations of the Trp-cage protein confirm this picture on the free energy landscape. Our analytical and computational results are consistent with a model in which the length and time scales of protein folding, as well as the modular nature of large proteins, are dictated primarily by inherent physical forces, whereas natural selection determines the native state.

Background: Direct identification of second systolic peaks of peripheral upper limb pulses (pSBP2) has been used to represent central systolic blood pressure (cSBP), but its accuracy at low SBP was questioned., Objectives: We investigated the relationship of pSBP2 with characteristics of central pressure waveforms., Methods: High-fidelity central aortic and right brachial pressure waveforms were simultaneously recorded using a custom-made dual pressure sensor catheter in 78 patients (65.9 ± 12.9 years) during catheterization for 285 measurements., Results: Overall agreement between cSBP and pSBP2 was good (mean difference -0.9 ± 4.8, r = 0.98), with a systematic bias at low SBP. We examined agreements of different waveform types according to the relationship of the second systolic peak of aortic pressure waveforms (cSBP2) to cSBP. Of type A (positive late systolic augmentation) and type B (zero augmentation) aortic pressure waveforms, in which cSBP = cSBP2, agreement between pSBP2 and cSBP was excellent (mean difference -0.4 ± 4.1, r = 0.99). There were 40 type C aortic pressure waveforms (negative augmentation; cSBP > cSBP2) with cSBP 107.2 ± 13.9 mmHg. Their cSBP2, compared with cSBP, showed closer agreement (mean difference -0.6 ± 3.2 vs. -4.0 ± 7.2 mmHg) and better correlation (r = 0.97 vs. 0.85, P = 0.03) with pSBP2., Conclusion: pSBP2 can be used with type A and B aortic pressure waveforms for estimation of cSBP. However, it should not be used with type C aortic pressure waveforms, typically at low SBP, because pSBP2 is closer to cSBP2 than cSBP. This explains why pSBP2 underestimates cSBP at low SBP.

Tuberculous aortitis is a rare disease entity indicative of disseminated tuberculosis. We report a case who presented with newly diagnosed hypertension one year after her last admission for pulmonary and pericardial tuberculosis. Chest CT and MRA, performed due to marked differences in pulse amplitude between upper and lower limbs, demonstrated long and severe segmental stenosis of the thoracic aorta. After the operation of bypass graft for the thoracic aorta, the patient recovered uneventfully. Tuberculous aortitis should be included in the list of differential diagnosis for secondary hypertension, especially if the patient has a recent disease history of pulmonary tuberculosis.

Among the macromolecular patterns of biological significance, right-handed α-helices are perhaps the most abundant structural motifs. Here, guided by experimental findings, we discuss both ultrafast initial steps and longer-time-scale structural dynamics of helix-coil transitions induced by a range of temperature jumps in large, isolated macromolecular ensembles of an α-helical protein segment thymosin β(9) (Tβ(9)), and elucidate the comprehensive picture of (un)folding. In continuation of an earlier theoretical work from this laboratory that utilized a simplistic structure-scrambling algorithm combined with a variety of self-avoidance thresholds to approximately model helix-coil transitions in Tβ(9), in the present contribution we focus on the actual dynamics of unfolding as obtained from massively distributed ensemble-convergent MD simulations which provide an unprecedented scope of information on the nature of transient macromolecular structures, and with atomic-scale spatiotemporal resolution. In addition to the use of radial distribution functions of ultrafast electron diffraction (UED) simulations in gaining an insight into the elementary steps of conformational interconversions, we also investigate the structural dynamics of the protein via the native (α-helical) hydrogen bonding contact metric which is an intuitive coarse graining approach. Importantly, the decay of α-helical motifs and the (globular) conformational annealing in Tβ(9) occur consecutively or competitively, depending on the magnitude of temperature jump.

As the simplest and most prevalent motif of protein folding, α-helix initiation is the starting point of macromolecular complexity. In this work, helix initiation was directly measured via ultrafast temperature-jump spectroscopy on the smallest possible helix nucleus for which only the first turn is formed. The rate's dependence on sequence, length, and temperature reveals the fastest possible events in protein folding dynamics, and it was possible to separate the rate-limiting torsional (conformational) diffusion from the fast annealing of the helix. An analytic coarse-grained model for this process, which predicts the initiation rate as a function of temperature, confirms this picture. Moreover, the stipulations of the model were verified by ensemble-converging all-atom molecular dynamics simulations, which reproduced both the picosecond annealing and the nanosecond diffusion processes observed experimentally.

The Euclidean distance matrix for n distinct points in ℝ r is generically of rank r + 2. It is shown in this paper via a geometric argument that its nonnegative rank for the case r = 1 is generically n .

In the past decade or so, semi-definite programming (SDP) has emerged as a powerful tool capable of handling a remarkably wide range of problems. This article describes an innovative application of SDP techniques to quadratic inverse eigenvalue problems (QIEPs). The notion of QIEPs is of fundamental importance because its ultimate goal of constructing or updating a vibration system from some observed or desirable dynamical behaviors while respecting some inherent feasibility constraints well suits many engineering applications. Thus far, however, QIEPs have remained challenging both theoretically and computationally due to the great variations of structural constraints that must be addressed. Of notable interest and significance are the uniformity and the simplicity in the SDP formulation that solves effectively many otherwise very difficult QIEPs.

Optimal immunity to microorganisms depends upon the regulated death of clonally expanded effector cells and the survival of a cohort of cells that become memory cells. After activation of naive T cells, CD44, a widely expressed receptor for extracellular matrix components, is upregulated. High expression of CD44 remains on memory cells and despite its wide usage as a "memory marker," its function is unknown. Here we report that CD44 was essential for the generation of memory T helper 1 (Th1) cells by promoting effector cell survival. This dependency was not found in Th2, Th17, or CD8(+) T cells despite similar expression of CD44 and the absence of splice variants in all subsets. CD44 limited Fas-mediated death in Th1 cells and its ligation engaged the phosphoinositide 3-kinase-Akt kinase signaling pathway that regulates cell survival. The difference in CD44-regulated apoptosis resistance in T cell subpopulations has important implications in a broad spectrum of diseases., (Copyright 2010 Elsevier Inc. All rights reserved.)

The advanced fabrication and surface engineering of superparamagnetic iron oxide nanoparticles (SPIONs) could offer excellent physiochemical features for noninvasive tumor imaging and drug delivery. The key issues of realization of maximized selective cancer targeting of SPIONs are minimization of uptake by macrophages, preferential binding to cancerous cells over neighboring normal cells, visualization of tumor cells prior to and after treatment and triggered drug release into target cells in a controlled fashion. In this article, we summarize the current status of fabrication of multifunctional SPION-based nanodevices specially designed for cancer-oriented diagnosis and therapy, with a focus on potential malignancy-targeting ligands' identification and development as nanocarriers. A number of examples of passive and active targeting strategies--lymphoangiogenesis markers, cellular metabolite receptors, extracellular matrix component receptors, neuropeptide receptors and receptor-mediated bypass of the blood-brain barrier--are described in detail.

Of special interest in molecular biology is the study of structural and conformational changes which are free of the additional effects of the environment. In the present contribution, we report on the ultrafast unfolding dynamics of a large DNA macromolecular ensemble in vacuo for a number of temperature jumps, and make a comparison with the unfolding dynamics of the DNA in aqueous solution. A number of coarse-graining approaches, such as kinetic intermediate structure (KIS) model and ensemble-averaged radial distribution functions, are used to account for the transitional dynamics of the DNA without sacrificing the structural resolution. The studied ensembles of DNA macromolecules were generated using distributed molecular dynamics (MD) simulations, and the ensemble convergence was ensured by monitoring the ensemble-averaged radial distribution functions and KIS unfolding trajectories. Because the order-disorder transition in free DNA implies unzipping, coiling, and strand-separation processes which occur consecutively or competitively depending on the initial and final temperature of the ensemble, DNA order-disorder transition in vacuo cannot be described as a two-state (un)folding process.

Quadratic matrix polynomials are fundamental to vibration analysis. Because of the predetermined interconnectivity among the constituent elements and the mandatory nonnegativity of the physical parameters, most given vibration systems will impose some inherent structure on the coefficients of the corresponding quadratic matrix polynomials. In the inverse problem of reconstructing a vibration system from its observed or desirable dynamical behavior, respecting the intrinsic structure becomes important and challenging both theoretically and practically. The issue of whether a structured inverse eigenvalue problem is solvable is problem dependent and has to be addressed structure by structure. In an earlier work, physical systems that can be modeled under the paradigm of a serially linked mass-spring system have been considered via specifically formulated inequality systems. In this paper, the framework is generalized to arbitrary generally linked systems. In particular, given any configuration of interconnectivity in a mass-spring system, this paper presents a mechanism that systematically and automatically generates a corresponding inequality system. A numerical approach is proposed to determine whether the inverse problem is solvable and, if it is so, computes the coefficient matrices while providing an estimate of the residual error. The most important feature of this approach is that it is problem independent, that is, the approach is general and robust for any kind of physical configuration. The ideas discussed in this paper have been implemented into a software package by which some numerical experiments are reported.

Protein structural integrity and flexibility are intimately tied to solvation. Here, we examine the effect that changes in bulk and local solvent properties have on protein structure and stability. We observe the change in solvation of an unfolding of the protein model, melittin, in the presence of a denaturant, trifluoroethanol. The peptide system displays a well defined transition in that the tetramer unfolds without disrupting the secondary or tertiary structure. In the absence of local structural perturbation, we are able to reveal exclusively the role of solvation dynamics in protein structure stabilization and the (un)folding pathway. A sudden retardation in solvent dynamics, which is coupled to the change in protein structure, is observed at a critical trifluoroethanol concentration. The large amplitude conformational changes are regulated by the local solvent hydrophobicity and bulk solvent viscosity.

In this article we consider consequences of spatial coherences and conformations in diffraction of (macro)molecules with different potential energy landscapes. The emphasis is on using this understanding to extract structural and temporal information from diffraction experiments. The theoretical analysis of structural interconversions spans an increased range of complexity, from small hydrocarbons to proteins. For each molecule considered, we construct the potential energy landscape and assess the characteristic conformational states available. For molecules that are quasiharmonic in the vicinity of energy minima, we find that the distinct conformer model is sufficient even at high temperatures. If, however, the energy surface is either locally flat around the minima or the molecule includes many degrees of conformational freedom, a Boltzmann ensemble must be used, in what we define as the pseudoconformer approach, to reproduce the diffraction. For macromolecules with numerous energy minima, the ensemble of hundreds of structures is considered, but we also utilize the concept of the persistence length to provide information on orientational coherence and its use to assess the degree of resonance contribution to diffraction. It is shown that the erosion of the resonant features in diffraction which are characteristic of some quasiperiodic structural motifs can be exploited in experimental studies of conformational interconversions triggered by a laser-induced temperature jump.

Superparamagnetic iron oxide nanoparticles (SPIONs) have attract a great deal of interest in biomedical research and clinical applications over the past decades. Taking advantage the fact that SPIONs only exhibit magnetic properties in the presence of an applied magnetic field, they have been used in both in vitro magnetic separation and in vivo applications such as hyperthermia (HT), magnetic drug targeting (MDT), magnetic resonance imaging (MRI), gene delivery (GD) and nanomedicine. Successful applications of SPIONs rely on precise control of the particle's shape, size, and size distribution and several synthetic routes for preparing SPIONs have been explored. Tailored surface properties specifically designed for cell targeting are often required, although the generic strategy involves creating biocompatible polymeric or non-polymeric coating and subsequent conjugation of bioactive molecules. In this review article, synthetic routes, surface modification and functionaliztion of SPIONs, as well as the major biomedical applications are summarized, with emphasis on in vivo applications.

A 2D free-energy landscape model is presented to describe the (un)folding transition of DNA/RNA hairpins, together with molecular dynamics simulations and experimental findings. The dependence of the (un)folding transition on the stem sequence and the loop length is shown in the enthalpic and entropic contributions to the free energy. Intermediate structures are well defined by the two coordinates of the landscape during (un)zipping. Both the free-energy landscape model and the extensive molecular dynamics simulations totaling over 10 mus predict the existence of temperature-dependent kinetic intermediate states during hairpin (un)zipping and provide the theoretical description of recent ultrafast temperature-jump studies which indicate that hairpin (un)zipping is, in general, not a two-state process. The model allows for lucid prediction of the collapsed state(s) in simple 2D space and we term it the kinetic intermediate structure (KIS) model.

Type 1 diabetes is a CD4 cell-dependent disease that results from destruction of insulin-producing beta cells in pancreatic islets. An ideal therapy would reverse diabetes shortly after onset when islet function in not yet fully ablated, and also prevent re-emergence of disease through the generation of memory cells that control the autoimmune response. In this study, we show that adaptive/induced polyclonal regulatory (TR) cells, which contain islet-reactive cells, fulfill these criteria in the NOD mouse model. CD4 cells induced to express FoxP3, IL-10, and TGF-beta1 in response to TCR signaling and TGF-beta1 can reverse diabetes with clinical restoration of prediabetic serum levels of IL-10. Unlike naturally occurring TR cells, these adaptive TR cells persist indefinitely (>1 year) as FoxP3(+), CD25(-) memory cells that self-renew. Establishment of memory is accompanied by narrowing of the T cell repertoire to usage of a single TCR beta-chain, Vbeta11, implying selection by Ag. With islet-specific adaptive TR cells, we show that memory is functionally stable and transferable. Therefore, adaptive TR cells, which can be readily generated from normal CD4 populations and become focused by Ag with induction of memory, may provide a treatment and a vaccine for the long-term cure of diabetes making them attractive as immunotherapeutic agents.

Herbal and other dietary supplements are popular among patients. Whether and how to establish and manage an herbal formulary remains a challenge for conventional managed-care organizations. Pharmacists and clinicians depend on evidence-based guidelines to help determine which products (whether pharmaceutical or herbal) to make available to patients. Evidence from randomized controlled trials that supports the use of most herbal supplements is scarce, yet for some supplements, credible evidence supports the possibility of efficacy. Quality control remains of concern for the supplement industry and for patients and clinicians considering the use of supplements.Clinicians may improve care by both disseminating educational materials and making available to patients popular quality-controlled herbal supplements deemed safe and effective. Kaiser Permanente Northwest (KPNW) has adopted a comprehensive and systematic approach to managing and stocking herbal supplements that can serve as a model for other conventional and managed care organizations.KPNW's dedicated Natural Products Advisory Committee (NPAC) has made considerable progress toward developing a constructive response to KPNW patients' herbal supplement use. Making supplements available at pharmacies can improve clinical outcomes, patient convenience, and quality control. NPAC currently limits its reviews to randomized, controlled trials and data from meta-analyses and systematic reviews for single-ingredient supplements. As interest in this area maintains steady growth, NPAC will continue to study how best to meet patients' needs.

Uptake 14CO2 from air and accumulation of 14C in the Chinese cabbage were studied by using the isotope-tracer techniques in order to get a better understanding of the environmental behavior of 14CO. The results show that the 14CO2, which was uptaken from the air by the leaf photosynthesis, would be sent to the other part of the Chinese cabbage and form a accumulating trend. The specific activity of 14C in every tissue increases linearly with time, and the increasing rate is in the range of 95.3 - 270.2 Bq/(g x d). The order of rate in tissues is as below: cabbage heart > leaf blade > leaf stalk > basal plate > root. The Chinese cabbage has an intensive concentration of 14CO2, and the concentration factor values increase rapidly with time. The concentration factor values in the cabbage heart reach the highest level (156.4 in 48 days) and following with the leaf blade (135.6 in 48 days). The specific activity in each part of the Chinese cabbage also increases with the introduction times of the 14CO2. Regression analysis has proved that the specific activity (C) of every part is positively correlated to the introducing times (N). Owing to its high accumulation of 14CO2, Chinese cabbage can be used as the indicator plant in the supervisor of 14CO2 air pollution.

The Ig kappa intronic and 3'kappa light chain enhancers have been shown to be necessary for V region hypermutation in kappa light chain transgenes. To investigate the role of the E mu intronic enhancer in V region hypermutation of heavy chain genes, E mu and its associated matrix attachment regions (MAR) were replaced with the SV40 or the cytomegalovirus (CMV) enhancer in a gamma2a construct that hypermutates its rearranged VDJ region in the NSO plasmacytoma and 18.81 pre-B cell lines. In this model in vitro system, mutation rates of stable transfectants were determined by reversion analysis using a V region stop codon that, when mutated, allowed the detection of cellular revertants by ELISA spot assay. The gamma2a constructs with the E mu, SV40 and CMV enhancers mutated at comparably high rates in the B cell lines, but not in L cells, indicating that the E mu enhancer and its associated MAR were not specifically required for IgH hypermutation in this system. In parallel experiments, the addition of the 3'alpha heavy chain enhancer (hs 1,2) or the 3'kappa light chain enhancer did not increase the mutation rate of a related mu reporter construct in which the associated VDJ mutates at a moderately low rate in NSO cells or in cell hybrids made between 18.81 and NSO. These results imply that cis-acting IgH elements that promote hypermutation may not be restricted to Ig-specific transcriptional enhancers.

Studies of endogenous and engineered Ig genes in mice have begun to reveal some of the cis-acting regions that are involved in the somatic hypermutation of variable regions in vivo. These studies suggest that the initiation of transcription plays a role in this process. However, it will be difficult to identify and manipulate the individual genetic elements and the trans-acting proteins that regulate and target the mutational events using solely in vivo assays. These studies would be greatly facilitated if constructs containing the genetic elements that are essential for V-region mutation could be transfected into cultured cells and undergo high rates of V-region mutation in vitro, and if permissive and non-permissive cell lines could be identified. Such in vitro systems would also allow a detailed molecular and biochemical analysis of this process. Here, we discuss some of the in vitro systems that have been developed and use data from our own studies in cultured cells to illustrate the potential benefits of studying V-region hypermutation in model in vitro systems.

In the course of an immune response, antibodies undergo affinity maturation in order to increase their efficiency in neutralizing foreign invaders. Affinity maturation occurs by the introduction of multiple point mutations in the variable region gene that encodes the antigen binding site. This somatic hypermutation is restricted to immunoglobulin genes and occurs at very high rates. The precise molecular basis of this process remains obscure. However, recent studies using a variety of in vivo and in vitro systems have revealed important regulatory regions, base motifs that are preferred targets of mutation and evidence that transcription may play an active role in hypermutation.

The variable (V) regions of immunoglobulin heavy and light chains undergo high rates of somatic mutation during the immune response. Although point mutations accumulate throughout the V regions and their immediate flanking sequences, analysis of large numbers of mutations that have arisen in vivo reveal that the triplet AGC appears to be most susceptible to mutation. We have stably transfected B cell lines with gamma2a heavy chain constructs containing TAG nonsense codons in their V regions that are part of either a putative (T)AGC hot spot or a (T)AGA non-hot spot motif. Using an ELISA spot assay to detect revertants and fluctuation analysis to determine rates of mutation, the rate of reversion of the TAG nonsense codon has been determined for different motifs in different parts of the V region. In the NSO plasma cell line, the (T)AGC hot spot motif mutates at rates of approximately 6 x 10(-4)/bp per generation and approximately 3 x 10(-5)/bp per generation at residues 38 and 94 in the V region. At each of these locations, the (T)AGC hot spot motif is 20-30 times more likely to undergo mutation than the (T)AGA non-hot spot motif. Moreover, the AGA non-hot spot motif mutates at as high a rate as the hot spot motif when it is located adjacent to hot spot motifs, suggesting that more extended sequences influence susceptibility to mutation.

There recently has been increasing interest at substance abuse treatment centers in smoking cessation treatment. Because a history of depression has been shown in other populations to complicate cessation efforts, the relationship between depression and nicotine dependence was tested in 726 methadone patients. Elevated odds of nicotine dependence given depression were found with three of four depression measures. Additional research is recommended to determine whether smoking cessation treatment will be more successful for methadone patients with a history of depression if it also addresses depression.

A subset of antiandrogen compounds, the N-aryl-3,3,3-trifluoro-2-hydroxy-2-methylpropanamides 1, were found to activate ATP sensitive potassium channels (KATP) and represent a new class of potassium channel openers (PCOs). A structure-activity relationship was carried out on the western region of this series with the goal of obtaining an activator of the ATP sensitive potassium channel suitable for use in the treatment of urge urinary incontinence. In particular three large 4-(N-aryl) substituents, the (N-phenyl-N-methylamino)sulfonyl, benzoyl, and 4-pyridylsulfonyl moieties, yielded non-antiandrogen, KATP potassium channel openers (39, 41, and 64, respectively) that are bladder selective in an in vivo rat model that simultaneously measures bladder contractions, heart rate, and blood pressure. Substitutions of the aryl rings of 41 and 64 gave several derivatives that also display selectivity in the in vivo rat model; however, none appear to offer a substantial advantage over 41 and 64. The PCO activity of 41 and 64 resides in the (S)-(-) enantiomers. ZD6169, 41(S), has been selected into development for the treatment of urge urinary incontinence.