April L. Harkins

Chitosan–cellulose composite for wound dressing material. Part 2. Antimicrobial activity, blood absorption ability, and biocompatibility

Chitosan (CS), a polysaccharide derived from chitin, the second most abundant polysaccharide, is widely used in the medical world because of its natural and nontoxic properties and its innate ability for antibacterial and hemostasis effects. In this study, the novel composites containing CS and cellulose (CEL) (i.e., [CEL + CS]), which we have previously synthesized using a green and totally recyclable method, were investigated for their antimicrobial activity, absorption of anticoagulated whole blood, anti-inflammatory activity through the reduction of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), and the biocompatibility with human fibroblasts. The [CEL + CS] composites were found to inhibit the growth of both Gram positive and negative micro-organisms. For examples, the regenerated 100% lyophilized chitosan material was found to reduce growth of Escherichia coli (ATCC 8739 and vancomycin resistant Enterococcus faecalis (ATCC 51299) by 78, 36, and 64%, respectively. The composites are nontoxic to fibroblasts; that is, fibroblasts, which are critical to the formation of connective tissue matrix were found to grow and proliferate in the presence of the composites. They effectively absorb blood, and at the same rate and volume as commercially available wound dressings. The composites, in both air-dried and lyophilized forms, significantly inhibit the production of TNF-α and IL-6 by stimulated macrophages. These results clearly indicate that the biodegradable, biocompatible and nontoxic [CEL + CS] composites, particularly those dried by lyophilizing, can be effectively used as a material in wound dressings.

Pain Perception after Isometric Exercise in Women with Fibromyalgia

OBJECTIVE The purpose of this study was to identify exercise protocols incorporating isometric contractions that provide pain relief in women with fibromyalgia. DESIGN A before-after trial. SETTING A physical therapy department in an academic setting. PARTICIPANTS Fifteen women (mean ± SD, 52±11y) with fibromyalgia. INTERVENTIONS Subjects completed 4 sessions: 1 familiarization and 3 experimental. The following randomized experimental sessions involved the performance of isometric contractions with the elbow flexor muscles that varied in intensity and duration: (1) 3 maximal voluntary contractions (MVCs), (2) 25% MVC held to task failure, and (3) 25% MVC held for 2 minutes. MAIN OUTCOME MEASURES Experimental pain (pain threshold and pain rating), Fibromyalgia Impact Questionnaire, and fibromyalgia pain intensity (visual analog scale). RESULTS After all 3 isometric contractions, there was considerable variability between subjects in the pain response. Based on the changes in experimental pain, subjects were divided into 3 groups (increase, decrease, no change in pain). Multiple regression analysis revealed that age, baseline experimental pain, and change in fibromyalgia pain intensity were significant predictors of the experimental pain response after the isometric contractions. CONCLUSIONS We identified subgroups of women with fibromyalgia based on how they perceived pain after isometric contractions. The greatest pain relief for women with fibromyalgia occurred at a younger age and in women with the greatest experimental pain before exercise. Additionally, we established a link between experimental and clinical pain relief after the performance of isometric contractions.

Supraspinal Fatigue Is Similar in Men and Women for a Low-Force Fatiguing Contraction

PURPOSE This study determined the contribution of supraspinal fatigue to the sex difference in neuromuscular fatigue for a low-intensity fatiguing contraction. Because women have greater motor responses to arousal than men, we also examined whether cortical and motor nerve stimulation, techniques used to quantify central fatigue, would alter the sex difference in muscle fatigue. METHODS In study 1, cortical stimulation was elicited during maximal voluntary contractions (MVC) before and after a submaximal isometric contraction at 20% MVC with the elbow flexor muscles in 29 young adults (20 ± 2.6 yr, 14 men). In study 2, 10 men and 10 women (19.1 ± 2.9 yr) performed a fatiguing contraction in the presence and absence of cortical and motor nerve stimulation. RESULTS Study 1: Men had a briefer time to task failure than women (P = 0.009). Voluntary activation was reduced after the fatiguing contraction (P < 0.001) similarly for men and women. Motor-evoked potential area and the EMG silent period increased similarly with fatigue for both sexes. Peak relaxation rates, however, were greater for men than women and were associated with time to task failure (P < 0.05). Force fluctuations, RPE, HR, and mean arterial pressure increased at a greater rate for men than for women during the fatiguing contraction (P < 0.05). Study 2: Time to task failure, force fluctuations, and all other physiological variables assessed were similar for the control session and stimulation session (P > 0.05) for both men and women. CONCLUSIONS Supraspinal fatigue was similar for men and women after the low-force fatiguing contraction, and the sex difference in muscle fatigue was associated with peripheral mechanisms. Furthermore, supraspinal fatigue can be quantified in both men and women without influencing motor performance.

Anxiety and stress can predict pain perception following a cognitive stress

Hoeger Bement, M.K., A. Weyer, M. Keller, A. Harkins, and S.K. Hunter. Anxiety and stress can predict pain perception following a cognitive stressor. PHYSIOL BEHAV 000-000. The purpose of this study was to investigate the influence of a cognitive stressor on pain perception and determine individual characteristics that may predict the pain response. Twenty-five subjects participated in three sessions: one familiarization and two experimental. The experimental sessions involved measurement of pain perception before and after 1) mental math tasks (stressor session) and 2) quiet rest (control session). Pain threshold and ratings were assessed with a mechanical noxious stimulus. Changes in stress and anxiety were examined with self-reported and physiological measures including questionnaires, visual analogue scales, and salivary cortisol levels. During the control session, stress and anxiety decreased and pain reports remain unchanged. During the stressor session, stress and anxiety increased and pain reports were variable among subjects. Based on the pain response to mental math, subjects were divided into three groups (increase, decrease or no change in pain). The increase-pain group (n=8) had lower baseline stress and anxiety, lower baseline pain reports, and large anxiety response following the mental math. In contrast, the decrease-pain group (n=9) had higher baseline stress and anxiety levels, higher baseline pain reports, and a large increase in cortisol levels. Thus, the differential response in the changes in pain perception was related to anxiety and stress levels prior to and during the cognitive stressor, indicating that psychosocial characteristics can help determine the stress-induced pain response.

The role of the menstrual cycle phase in pain perception before and after an isometric fatiguing contraction

The purpose of this study was to compare exercise-induced analgesia in young women after a fatiguing isometric contraction during different phases of the menstrual cycle. Twenty female subjects performed a submaximal (25% maximal voluntary contraction) isometric contraction until task failure during both the mid-follicular and mid-luteal phases of their menstrual cycle. Pain perception (i.e., pain threshold and pain ratings) was measured before and after the isometric fatiguing contraction. Other measures included mean arterial pressure, heart rate, and anxiety levels. Time to task failure of the fatiguing contraction was similar for the two phases of the menstrual cycle. Following the performance of the isometric contraction: (1) pain thresholds increased and pain ratings decreased; (2) anxiety levels increased; and (3) mean arterial pressure and heart rate increased. These changes were not dependent on the phase of the menstrual cycle. Thus, the menstrual cycle phase does not influence the magnitude of exercise-induced analgesia.

Recyclable synthesis, characterization, and antimicrobial activity of chitosan-based polysaccharide composite materials

We have successfully developed a simple and totally recyclable method to synthesize novel, biocompatible, and biodegradable composite materials from cellulose (CEL) and chitosan (CS). In this method, [BMIm(+) Cl(-) ], an ionic liquid (IL), was used as a green solvent to dissolve and synthesize the [CEL+CS] composites. Since, the IL can be removed from the composites by washing them with water, and recovered by distilling the washed solution, the method is totally recyclable. Spectroscopic and imaging techniques including XRD, FTIR, NIR, and SEM were used to monitor the dissolution, to characterize and to confirm that CEL and CS were successfully regenerated. More importantly, we have successfully demonstrated that [CEL+CS] composite is particularly suited for many applications including antimicrobial property. This is because the composites have combined advantages of their components, namely superior chemical and mechanical stability (from CEL) and bactericide (from CS). Results of tensile strength measurements clearly indicate that adding CEL into CS substantially increase its tensile strength. Up to 5× increase in tensile strength can be achieved by adding 80% of CEL into CS. Results of in vitro antibacterial assays confirm that CS retains its antibacterial property in the composite. More importantly, the composites reported here can inhibit growth of wider range of bacteria than other CS-based materials prepared by conventional methods; that is over 24 h period, the composites substantially inhibited growth of bacteria such as MRSA, VRE, S. aureus, E. coli. These are bacteria that are often found to have the highest morbidity and mortality associated with wound infections.

FACILE SYNTHESIS, CHARACTERIZATION AND ANTIMICROBIAL ACTIVITY OF CELLULOSE-CHITOSAN- HYDROXYAPATITE COMPOSITE MATERIAL, A POTENTIAL MATERIAL FOR BONE TISSUE ENGINEERING

Hydroxyapatite (HAp) is often used as a bone-implant material because it is biocompatible and osteoconductive. However, HAp possesses poor rheological properties and it is inactive against disease-causing microbes. To improve these properties, we developed a green method to synthesize multifunctional composites containing: (1) cellulose (CEL) to impart mechanical strength; (2) chitosan (CS) to induce antibacterial activity thereby maintaining a microbe-free wound site; and (3) HAp. In this method, CS and CEL were co-dissolved in an ionic liquid (IL) and then regenerated from water. HAp was subsequently formed in situ by alternately soaking [CEL+CS] composites in aqueous solutions of CaCl2 and Na2 HPO4 . At least 88% of IL used was recovered for reuse by distilling the aqueous washings of [CEL+CS]. The composites were characterized using FTIR, XRD, and SEM. These composites retained the desirable properties of their constituents. For example, the tensile strength of the composites was enhanced 1.9 times by increasing CEL loading from 20% to 80%. Incorporating CS in the composites resulted in composites which inhibited the growth of both Gram positive (MRSA, S. aureus and VRE) and Gram negative (E. coli and P. aeruginosa) bacteria. These findings highlight the potential use of [CEL+CS+HAp] composites as scaffolds in bone tissue engineering.

Molecular Diagnosis of Sexually Transmitted Chlamydia trachomatis in the United States

Chlamydia, with its Chlamydia trachomatis etiology, is the most common bacterial sexually transmitted infection in the United States and is often transmitted via asymptomatic individuals. This review summarizes traditional and molecular-based diagnostic modalities specific to C. trachomatis. Several commercially available, FDA-approved molecular methods to diagnose urogenital C. trachomatis infection include nucleic acid hybridization, signal amplification, polymerase chain reaction, strand displacement amplification, and transcription-mediated amplification. Molecular-based methods are rapid and reliable genital specimen screening measures, especially when applied to areas of high disease prevalence. However, clinical and analytical sensitivity for some commercial systems decreases dramatically when testing urine samples. In vitro experiments and clinical data suggest that transcription-mediated amplification has greater analytical sensitivity than the other molecular-based methods currently available. This difference may be further exhibited in testing of extragenital specimens from at-risk patient demographics. The development of future molecular testing could address conundrums associated with confirmatory testing, medicolegal testing, and test of cure.

Effect of Preanalytical Processing of ThinPrep Specimens on Detection of High-Risk Human Papillomavirus by the Aptima HPV Assay

ABSTRACT Two important preanalytical protocols performed on liquid-based cytological specimens, namely, automated cytology processing and glacial acetic acid (GAA) treatment, may occur prior to the arrival of specimens in a molecular diagnostics laboratory. Ninety-two ThinPrep vials previously positive for high-risk human papillomavirus (HPV) via the Cervista HPV HR test were preselected and alternated with 92 previously negative ThinPrep vials. The specimen set was processed in a consecutive fashion by an automated cytology processor without fastidious decontamination precautions. Carryover potential was subsequently assessed by performance of the Aptima HPV assay on aliquots from reprocessed ThinPrep vials. All previously negative ThinPrep vials yielded a negative result following routine automated cytology processing, despite close proximity to known-positive ThinPrep vials. In separate experiments, aliquots from 236 ThinPrep vials were forwarded for tandem analysis with and without GAA treatment. Data from GAA- and mock-treated specimens generated by Aptima HPV were compared to correlate data generated by Cervista. A 99.2% concordance of Aptima HPV results from GAA-treated and mock-treated specimens was noted. This result differed from the concordance result derived from Cervista (91.5%; P < 0.0002). Of the initially positive Cervista results, 21.9% reverted to negative following GAA treatment; the correlate value was 2.7% for Aptima HPV (P = 0.01). While deleterious effects of GAA treatment on genomic DNA were noted with Cervista (P = 0.0015), GAA treatment had no significant effects on Aptima HPV specimen signal/cutoff ratios or amplification of internal control RNA (P ≥ 0.07). The validity of an Aptima HPV result is independent of GAA treatment and routine automated cytology processing.

An oleate‐stimulated, phosphatidylinositol 4,5‐bisphosphate‐independent phospholipase D in Schizosaccharomyces pombe

Phospholipase D1 (PLD1) is an important enzyme involved in lipid-mediated signal transduction and membrane dynamics in eukaryotes. PLD1 preferentially hydrolyzes phosphatidylcholine to phosphatidic acid. This potent second messenger is involved in cytoskeletal reorganization, secretion, and membrane trafficking in eukaryotic cells. In Saccharomyces cerevisiae, PLD1 is involved in polarized growth and morphogenesis during pheromone response and sporulation. The presence of a PLD activity in Schizosaccharomyces pombe is demonstrated. PLD activity was able to hydrolyze a fluorescently labeled analog of phosphatidylcholine and was capable of performing the transphosphatidylation reaction characteristic of PLDs. Schizosaccharomyces pombe PLD activity was unaffected by phosphatidylinositol 4,5 bisphosphate (PIP(2)), but was slightly stimulated by oleate. PLD activity was shown to increase when the S. pombe cells underwent mating and sporulation. Here, we also report the molecular cloning of the first phospholipase D isoform from an S. pombe genomic DNA library (EMBL accession no. FN547388). Comparisons of three divergent yeasts, S. pombe, S. cerevisiae, and Candida albicans, with respect to the PLD enzymes revealed differences in regulation by oleate and PIP(2). Even with high homology in the protein sequences between the PLD1 enzymes of S. cerevisiae, C. albicans, and S. pombe, there was variation with the effects of the regulators.