Osseointegration for Lower Limb Amputation: Understanding the Risk Factors and Time Courses of Soft Tissue Complications.

PURPOSE: Lower-limb osseointegrated prostheses are a novel alternative to traditional socket-suspended prostheses, which are often associated with poor fit, soft tissue damage, and pain. Osseointegration eliminates the socket-skin interface and allows for weight-bearing directly on the skeletal system. However, these prostheses can also be complicated by postoperative issues that can negatively impact mobility and quality of life. Little is known about the incidence of or risk factors for these complications as few centers currently perform the procedure. METHODS: A retrospective analysis was performed on all patients who underwent single-stage lower limb osseointegration at our institution between 2017 and 2021. Patient demographics, medical history, operative data, and outcomes were collected. Fisher exact test and unpaired t tests were performed to identify risk factors for each adverse outcome, and time-to-event survival curves were generated. RESULTS: Sixty patients met our study criteria: 42 males and 18 females with 35 transfemoral and 25 transtibial amputations. The cohort had an average age of 48 years (range, 25-70 years) and follow-up period of 22 months (range, 6-47 months). Indications for amputation were trauma (50), prior surgical complication (5), cancer (4), and infection (1). Postoperatively, 25 patients developed soft tissue infections, 5 developed osteomyelitis, 6 had symptomatic neuromas, and 7 required soft tissue revisions. Soft tissue infections were positively correlated with obesity and female sex. Neuroma development was associated with increased age at osseointegration. Neuromas and osteomyelitis were both associated with decreased center experience. Subgroup analysis by amputation etiology and anatomic location did not show significant differences in outcomes. Notably, hypertension (15), tobacco use (27), and prior site infection (23) did not correlate with worse outcomes. Forty-seven percent of soft tissue infections occurred in the 1 month after implantation, and 76% occurred in the first 4 months. CONCLUSIONS: These data provide preliminary insights into risk factors for postoperative complications arising from lower limb osseointegration. These factors are both modifiable (body mass index, center experience), and unmodifiable (sex, age). As this procedure continues to expand in popularity, such results are necessary to inform best practice guidelines and optimize outcomes. Further prospective studies are needed to confirm the above trends.

Exploring PCDD/Fs and potentially toxic elements in sewage sludge during smouldering treatment.

Potentially toxic elements (PTEs), persistent organic pollutants, and emerging contaminants make sewage sludge management challenging. There is significant interest in thermal treatment technologies that can destroy these compounds. The most common thermal treatment, incineration, poses risks due to formation and/or release of hazardous substances in process emissions such as polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs) and PTEs. Smouldering has been introduced recently as a potential treatment for managing sewage sludge. Smouldering systems present several advantages over traditional incinerators; however, there are still uncertainties regarding process by-products. This key question was investigated in three laboratory-scale tests (0.08 m radius) and five oil drum-scale tests (0.3 m radius) that were evaluated for PCDD/Fs and PTEs in the mixture before and after treatment as well as in process emissions. Volatile organic compounds (VOCs) were also measured. These experiments represent a broad spectrum of conditions to evaluate process emissions, from robust self-sustaining to extinction of smouldering. Robust smouldering had negligible PCDD/Fs in process emissions. Weak smouldering had low levels of PCDD/Fs (emissions factor: 3.3 ± 0.3 µg TEQ/Mg dried sludge destroyed), levels less than uncontrolled emissions from commercial incinerators. Overall, smouldering acted as a sink for PCDD/Fs, as only 0-3% of the PCDD/Fs originally present in the sludge were released in the emissions, and >99% of the remainder were destroyed with <1% remaining in post-treatment ash. No evidence was found to support de novo synthesis or precursor reactions forming new PCDD/Fs. In addition, 94-100% of all the PTEs analyzed were retained in the post-smouldered material. These results indicate that only minimal emissions treatment for PTEs, PCDD/Fs, and VOCs may be necessary for future sewage sludge smouldering systems. These low emissions risks combined with its unique ability to handle high moisture content waste, indicate that smouldering has significant potential as a valuable waste management technique.

Pharmacodynamics of ceftriaxone for the treatment of methicillin-susceptible Staphylococcus aureus: is it a viable treatment option?

Ceftriaxone is a broad-spectrum cephalosporin that may be one option to treat methicillin-susceptible Staphylococcus aureus (MSSA). Although MSSA may be susceptible to ceftriaxone, the minimum inhibitory concentration (MIC) is generally two- to four-fold higher than other susceptible bacterial pathogens. This study aimed to explore the pharmacodynamics of ceftriaxone against MSSA and to determine the likely optimal dose. A hollow-fibre infection model was used with one clinical MSSA isolate (MIC = 4 mg/L) at an initial inoculum of 1 × 106 CFU/mL. Ceftriaxone dosing regimens of 1 g once and twice daily and 2 g once and twice daily were simulated. Ceftriaxone 1 g dosing regimens did not substantially impact bacterial killing within the first 12 h. Conversely, when administered as a 2 g dose either once or twice daily, an approximate 1-log10 bacterial reduction was observed where it plateaued for up to 96 h. No resistance was identified. Only a high ceftriaxone dose of 2 g twice daily achieves bacterial killing and sustained inhibition of bacterial growth. Ceftriaxone at routinely used doses is unsuitable for the treatment of MSSA infections and alternative agents should be preferentially used.

Curcumin induced oxidative stress attenuation by N-acetylcysteine co-treatment: a fibroblast and epithelial cell in-vitro study in idiopathic pulmonary fibrosis.

BACKGROUND: Idiopathic Pulmonary Fibrosis (IPF) is a fatal lung disease of unknown etiology with only two federally approved drug options. Given the complex molecular pathogenesis of IPF involving multiple cell types and multiple pathways, we explore the effects of a potential antifibrotic and antioxidant drug combination. Curcumin is a polyphenolic compound derived from turmeric with significant biological activity including a potential antifibrotic capacity. N-acetylcysteine (NAC) is a precursor to the antioxidant glutathione. To advance our understanding of these molecules, and to identify a clinical application, we present a small number of focused experiments that interrogates the effect of curcumin and NAC on pathways relevant to IPF in both fibroblasts and epithelial cells. METHODS: Primary epithelial cell and fibroblasts isolated from patients with IPF were challenged with a combination treatment of NAC and curcumin. Evaluation of the antifibrotic potential and effect on oxidative stress was performed through QPCR gene expression analysis and functional assays including scratch tests, viability assays, and measurement of induced reactive oxygen species. RESULTS: We demonstrate that curcumin alone does have antifibrotic potential, but that effect is accompanied by proapoptotic increases in oxidative stress. Coupled with this, we find that NAC alone can reduce oxidative stress, but that epithelial cell viability is decreased through this treatment. However, co-administration of these two molecules decreases oxidative stress and maintains high cell viability in both cell types. In addition, this co-treatment maintains an antifibrotic potential. CONCLUSIONS: These findings suggest a novel application for these molecules in IPF and encourage further exploration of this potential therapeutic approach.

Perfusion alters free zinc levels in the rodent brain.

BACKGROUND: Fixation of brain tissue is a common practice which allows preservation of tissue and aids in preventing structural and chemical abnormalities. However, fixation procedures may disrupt the levels of biometals such as zinc when compared to tissue that is fresh-frozen. Thus, we sought to determine if any differences in free-zinc levels exist between perfused and fresh-frozen tissue. Zinc is an essential biometal critical for cellular communication and memory and exists in both bound and free forms; the latter playing critical roles in synaptic communication. New method: C57BL/6 J mice were divided into two water types: those given lab water and those given water supplemented with 10 ppm zinc carbonate. Perfusion was carried out with 4% paraformaldehyde on half of the animals in each water group to assess the impact on levels of free Zn as measured through Zinpyr-1 fluorescence. RESULTS: There were significant differences in Zn fluorescence values between Zn-supplemented and lab water groups as well as between perfused and fresh-frozen tissues in the dentate gyrus and CA3 regions of the hippocampus, regions critical in learning & memory. Comparison with existing methods: These results show that when determining a method for euthanasia, any future histological techniques involving assessment of metal content should first be considered. CONCLUSIONS: Researchers must be cautious with the way in which tissue is collected and treated since this can lead to misleading conclusions when linking changes in behavior and relative levels of trace metals.

Infant Parenteral Nutrition Remains a Significant Source for Aluminum Toxicity.

BACKGROUND: Aluminum toxicity is associated with anemia, impaired bone metabolism, neurologic defects, and parenteral nutrition (PN)-associated liver disease. This element is a ubiquitous contaminant of PN components, especially in infant formulations. We assessed the current levels of aluminum contamination in infant PN at a level III neonatal intensive care unit. MATERIALS AND METHODS: Thirty samples of PN prepared in the same hospital for infants aged <30 days (mean [SD] weight, 1.54 [0.71] kg) were collected from discarded solution. Each sample was analyzed for aluminum content via inductively coupled plasma mass spectrometry. The components of PN (from label) and measured aluminum content were then compared using linear regression and 1-way analysis of variance. RESULTS: The mean (SD) aluminum contamination of infant PN was 14.02 (6.51) mcg/kg/d. Only 3 samples were <5 mcg/kg/d. Aluminum levels and infant weight were not associated. Linear regression revealed a significant correlation between aluminum and both calcium gluconate ( P < .0001) and phosphate ( P = .05), with a trend between aluminum and potassium ( P = .07). CONCLUSIONS: Aluminum contamination in infant PN remains almost 3 times higher than the advised maximum exposure (<5 mcg/kg/d, Food and Drug Administration 2004). Unexpectedly, an association between infant weight and aluminum exposure was not apparent, likely due to the homogeneity of our population. Isolating the source of aluminum contamination is difficult, as multiple components appear to be involved. Calcium gluconate is likely still a major contributor, but further investigations into individual components are warranted to promote the reduction of aluminum in infant PN.

Phomopsolides and related compounds from the alga-associated fungus, Penicillium clavigerum.

For the past fifteen years we have studied the secondary metabolites of extremophilic fungi from the Berkeley Pit, an abandoned acid mine waste lake. Fungi associated with an acid-tolerant alga have also been harvested from the Pit. Penicillium clavigerum Demelius was isolated from the green alga Chlorella vulgaris Beyerinck [Beijerinck]. In culture it produced the known compounds phomfuranone (1), patulin (2), dimethylphthalides (3) and (4), phomopsolide A (5), phomopsolide C (6), phomopsolide B (7), phomopsolide E (8), phomopsolide F (9), and phompyrone (10) and the new compound berkbenzofuran thioester (11). Compounds 5 and 6 were potent inhibitors (IC50 < 10 microM) of specific and established human cancer cell lines.

In vitro screening of plant lectins and tropical plant extracts for anthelmintic properties.

Lectins are plant secondary metabolites (PSM) found in many forages and which may confer anthelmintic properties to gastrointestinal parasites through disrupting the development of parasitic larvae throughout its life cycle. In experiment 1, the ability of the plant lectins jacalin (JAC), concanavalin A (Con A), phytohemagglutinin E2L2 (PHA-E2L2), phytohemagglutinin L4 (PHA-L4), phytohemagglutinin E3L (PHA-E3L), kidney bean albumin (KBA), Robinia pseudoacacia agglutinin (RPA), Maackia amurensis lectin (MAA), Maclura pomifera agglutinin (MAA), Dolichos biflorus agglutinin (DBA), wheat germ agglutinin (WGA) and Galanthus nivalis agglutinin (GNA) to disrupt the feeding of the first stage larvae (L(1)) of the sheep gastro-intestinal nematodes (GIN) Teladorsagia circumcincta, Haemonchus contortus and Trichostrongylus colubriformis was investigated using a larval feeding inhibition test (LFIT). Only PHA-E3L, WGA and Con A had a potent effect on disrupting larval feeding of all of the three species of GIN investigated. The lectin concentration required to inhibit feeding in 50% of L(1) (IC50) was 7.3±1.2, 8.3±1.4 and 4.3±1.7 µg/ml for PHA-E3L; 59.1±32.4, 58.7±11.9 and 8.1±7.0 µg/ml for Con A and 78.9±11.2, 69.4±8.1 and 28.0±14.1 µg/ml for WGA for T. circumcincta, H. contortus and T. colubriformis larvae, respectively (P=0.006). The addition of the lectin inhibitors fetuin, glucose/mannose or N-acetylglucosamine for PHA-E3L, Con A and WGA, respectively, caused an increase in the proportion of larvae that had fed at all concentrations for PHA-E3L only. In experiment 2, the effect of extracts from the tropical plants Azadiractha indica, Trichanthera gigantea, Morus alba, Gliricidia sepium and Leucaena leucocephala on the feeding behaviour of H. contortus L(1,) was examined. A. indica, T. gigantea and M. alba failed to inhibit 50% of larvae from feeding at concentrations up to 10mg plant extract per ml. In contrast, both G. sepium and L. leucocephala demonstrated a dose-dependent effect on larval feeding with respective IC50 estimates (mean±s.e.) of 0.015 mg/ml ±0.001 and 3.465 mg/ml ±0.144, effects which were partly reversed by the inclusion of either the tannin inhibitor polyethylene glycol or the lectin inhibitor Fetuin. These studies demonstrate that plant lectins can have an inhibitory effect on the feeding behaviour of first stage larvae of ovine GIN in vitro. Moreover they also provide novel evidence that lectins may contribute to the anthelmintic properties of some tropical forage plant extracts, such as G. sepium and L. leucocephala.

Effect of dietary supplementation with glutamine and a combination of glutamine-arginine on intestinal health in twenty-five-day-old weaned rabbits.

The effect of dietary supplementation with 1% l-glutamine and a combination of 1% l-glutamine and 0.5% l-arginine on intestinal health was examined in weaned rabbits. A basal diet was formulated to meet nutrient recommendations. Another 2 diets were formulated by adding 1% (as-fed basis) Gln or a mixture of 1% (as-fed basis) Gln + 0.5% (as-fed basis) Arg (Gln-Arg) to the basal diet. In Exp. 1, a total of 357 rabbits were blocked by litter and assigned at random to the experimental diet to determine mortality (119 per diet) and growth performance (35 per diet; from weaning at 25 to 56 d of age). Rabbits were fed the experimental diets for a 2-wk period and thereafter received a commercial diet. Rabbits weaned at 25 d (blocked by litter and assigned at random to diets) were slaughtered at 35 d and used to determine apparent ileal digestibility of DM, CP, and AA (Exp. 2, a total of 60 rabbits), intestinal morphology, N-aminopeptidase and myeloperoxidase intestinal activity, the expression of PPARgamma at the ileum and kidney, serum immunoglobulin in healthy and sick rabbits (Exp. 3, a total of 24 rabbits), and ileal and cecal microbial composition by PCR-RFLP (Exp. 4, a total of 45 rabbits). Dietary treatment did not affect ADG, ADFI, or G:F, during the entire fattening period. Supplementation with Gln reduced mortality during the first 2 wk and the whole fattening period from 18.5 to 8.4% (P = 0.023) and from 31.9 to 20.2% (P = 0.039), respectively, whereas no effect was detected for Arg supplementation. Among all the variables studied, the reduction on mortality due to Gln was related to a reduced intestinal colonization (Eimeria lesions) and changes on microbial ecosystem in the ileum and cecum, reducing the frequency of detection of Clostridium spp. (from 86.7 to 33.3%, P = 0.003) at the ileum, and Helicobacter spp. at the ileum (from 86.7 to 46.7%, P = 0.003) and at the cecum (from 86.7 to 46.7, P = 0.028), whereas no effect was detected for Arg supplementation. In conclusion, 1% l-Gln supplementation to postweaned rabbit diets decreased fattening mortality and modified the intestinal microbiota (although no consistent effects were observed on mucosal histology or inflammatory and systemic immune response). Diets containing a combination of 1% Gln and 0.5% Arg were of little additional benefit.

Improvement of agreement in TCM diagnosis among TCM practitioners for persons with the conventional diagnosis of rheumatoid arthritis: effect of training.

OBJECTIVE: To investigate whether a training process that focused on consensus on Traditional Chinese Medicine (TCM) diagnostic criteria will improve the agreement of TCM diagnosis on patients with rheumatoid arthritis (RA). DESIGN: The design was a prospective survey. SETTING: The study was conducted at the General Clinical Research Center, University of Maryland Hospital System, Baltimore, MD. SUBJECTS: The participants were 42 patients with RA. PRACTITIONERS: The practitioners included 3 licensed acupuncturists with a minimum of 5 years' licensure and education in Chinese herbs. METHODS: A training session of TCM diagnostic procedures was conducted with an open case discussion and "real time" practice. After the training, 3 TCM practitioners examined the same 42 patients with RA separately. Patients filled out a questionnaire to serve as the data for the "Inquiry" component while physical examinations, including observations of tongue and palpation of radial pulse, were conducted by the 3 practitioners. Each practitioner provided a TCM diagnosis based upon the examination results. These diagnoses were then examined with respect to the rate of agreement among the 3 practitioners. RESULTS: The average agreement with respect to the TCM diagnoses among the 3 pairs of TCM practitioners was 73% (64.3%-85.7%). Statistically significant differences were found between this study and the two previous studies (p < 0.001). CONCLUSIONS: After training focused on consensus on TCM diagnostic criteria, we found that these 3 same TCM practitioners who were used in phase II of the study produced a significantly higher agreement when compared to study phase I or phase II. Our study suggests that improved consensus on TCM diagnostic criteria results in increased agreement of diagnosis.

Relationship between dietary-induced changes in intestinal commensal microflora and duodenojejunal myoelectric activity monitored by radiotelemetry in the rat in vivo.

Interdigestive intestinal motility, and especially phase III of the migrating myoelectric/motor complex (MMC), is responsible for intestinal clearance and plays an important role in prevention of bacterial overgrowth and translocation in the gut. Yet previous results from gnotobiotic rats have shown that intestinal microflora can themselves affect the characteristics of the myoelectric activity of the gut during the interdigestive state. Given that the composition of the intestinal microflora can be altered by dietary manipulations, we investigated the effect of supplementation of the diet with synbiotics on intestinal microflora structure and the duodenojejunal myoelectric activity in the rat. To reduce animal distress caused by restraint and handling, which can itself affect GI motility, we applied radiotelemetry for duodenojejunal EMG recordings in conscious, freely moving rats. Thirty 16-month-old Spraque-Dawley rats were used. The diet for 15 rats (E group) was supplemented with chicory inulin, Lactobacillus rhamnosus and Bifidobacterium lactis. The remaining 15 rats were fed control diet without supplements (C group). Three rats from each group were implanted with three bipolar electrodes positioned at 2, 14 and 28 cm distal to the pylorus. After recovery, two 6 h recordings of duodenojejunal EMG were carried out on each operated rat. Subsequently, group C rats received feed supplements and group E rats received only control diet for 1 week, and an additional two 6 h recordings were carried out on each of these rats. Non-operated C and E rats were killed and samples of GI tract were collected for microbiological analyses. Supplementation of the diet with the pro- and prebiotics mixture increased the number of bifidobacteria, whereas it decreased the number of enterobacteria in jejunum, ileum, caecum and colon. In both caecum and colon, the dietary supplementation increased the number of total anaerobes and lactobacilli. Treatment with synbiotics increased occurrence of phase III of the MMC at all three levels of the small intestine. The propagation velocity of phase III in the whole recording segment was also increased from 3.7 +/- 0.2 to 4.4 +/- 0.2 cm min(-1) by dietary treatment. Treatment with synbiotics increased the frequency of response potentials of the propagated phase III of the MMC at both levels of the jejunum, but not in the duodenum. In both parts of the jejunum, the supplementation of the diet significantly decreased the duration of phase II of the MMC, while it did not change the duration of phase I and phase III. Using the telemetry technique it was demonstrated that changes in the gastrointestinal microflora exhibited an intestinal motility response and, more importantly, that such changes can be initiated by the addition of synbiotics to the diet.

Involvement of the p97-Ufd1-Npl4 complex in the regulated endoplasmic reticulum-associated degradation of inositol 1,4,5-trisphosphate receptors.

Inositol 1,4,5-trisphosphate (IP(3)) receptors form tetrameric, IP(3)-gated channels in endoplasmic reticulum membranes that govern the release of Ca(2+) from this organelle. In response to activation of certain G protein-coupled receptors that persistently elevate IP(3) concentration, IP(3) receptors are ubiquitinated and degraded by the ubiquitin-proteasome pathway. IP(3) receptor ubiquitination is mediated by the ubiquitin-conjugating enzyme, (mam)Ubc7, a component of the endoplasmic reticulum-associated degradation pathway. However, the mechanism by which ubiquitinated IP(3) receptors are transferred to the proteasome is not known. Here, we examine this process and show in several mammalian cell types that the ATPase p97 associates with IP(3) receptors in response to hormonal stimuli that induce IP(3) receptor ubiquitination. To examine the functional relevance of the p97 interaction with IP(3) receptors, we stably and specifically reduced p97 protein levels by 62 +/- 3% in Rat-1 fibroblasts using RNA interference. In these cells, endothelin-1-induced IP(3) receptor degradation was markedly retarded and the accumulation of ubiquitinated IP(3) receptors was markedly enhanced. These effects were reversed by expression of exogenous p97. In addition, Ufd1 and Npl4, which complex with p97, also associated with IP(3) receptors upon hormonal stimulation. We conclude that the p97-Ufd1-Npl4 complex couples ubiquitinated IP(3) receptors to proteasomal degradation and, thus, plays a key role in IP(3) receptor processing. These data also establish that the p97-Ufd1-Npl4 complex mediates endoplasmic reticulum-associated degradation in mammalian cells.

Variability in the traditional Chinese medicine (TCM) diagnoses and herbal prescriptions provided by three TCM practitioners for 40 patients with rheumatoid arthritis.

OBJECTIVE: To ascertain if previous findings of low levels of agreement of Traditional Chinese Medicine (TCM) pattern diagnoses made by TCM practitioners in patients with rheumatoid arthritis (RA) were a function of practitioner differences or would be replicated with a different sample of clinicians, and to examine the relationship between TCM diagnosis and herbal treatment plans. DESIGN: A prospective survey. SETTING: General clinical research center, University of Maryland Hospital System, Baltimore, MD. SUBJECTS: Forty (40) patients with RA. PRACTITIONERS: Licensed acupuncturists with at least 5 years' experience and education in Chinese herbs. METHODS: Three (3) TCM practitioners examined the same 40 RA patients separately, following the traditional Four Diagnostic Methods. Patients filled out questionnaires and physical examinations, including observations of the tongue and palpation of radial pulse, were conducted by the 3 practitioners. Each practitioner then provided both a TCM diagnosis and an herbal prescription. These diagnoses/prescriptions were examined with respect to the rate of agreement among the 3 practitioners. RESULTS: The average agreement with respect to the TCM diagnoses among the 3 TCM practitioners was 31.7 % (range, 27.5-35%). The degree to which the herbal prescriptions agreed with textbook recommended practice for each TCM diagnosis was 91.7% (range, 85-100%). The most commonly used TCM assessments in arriving at these diagnoses were inquiry about factors affecting pain and pulse diagnosis. No statistically significant differences were found between this study and our previous study regarding the level of agreement on TCM diagnosis. CONCLUSION: The average agreement of the diagnoses provided by 3 TCM practitioners was at the same low level as previously reported. No association was found between the diagnostic methods used and the consistency of diagnosis. Both studies, however, found a high degree of consistency between the TCM pattern diagnoses provided and the herbal treatment plans made as a result of those diagnoses.

Involvement of peripheral opioid mechanisms in electroacupuncture analgesia.

The involvement of the peripheral opioid system in modulating inflammatory pain has been well documented. This study aimed to investigate the possibility of electroacupuncture (EA)-mediated peripheral opioid release. Rats were injected with complete Freund's adjuvant in one of the hind paws to induce localized inflammatory pain. The pain behavioral changes were measured by paw withdrawal latency (PWL) to a noxious thermal stimulus. At day 5 of inflammation, rats received a second injection of saline or opioid antagonists into the inflamed paw, followed by EA at 30 Hz, 2 mA, and 0.1 ms for 30 minutes. The EA was conducted at acupuncture point GB30. A control was used in which needles were inserted at GB30 but no electrical stimulation was applied. Rats receiving EA showed a significantly longer PWL as compared with the control from 30 minutes to three hours after EA treatment. Intraplantar but not intraperitoneal injection of naloxone methiodide, a peripherally acting opioid receptor antagonist, eliminated the analgesic effect at 30 minutes after EA treatment. Intraplantar injection of an antibody against beta-endorphin and a corticotropin-releasing factor antagonist also produced a reduction in PWL in rats receiving EA. These data strongly suggest that peripheral opioids are released by EA at the inflammatory site.

Immunological implications of the use of plant lectins for drug and vaccine targeting to the gastrointestinal tract.

Plant lectins are under consideration as targeting agents to enhance the efficacy of orally administered drugs and vaccines. A significant issue that must be considered is the immunogenicity of these molecules since an immune response to the targeting agent may interfere with its ability to interact with the epithelium. In contrast, the ability of certain lectins to activate the immune system may be exploited in the delivery of vaccines. We previously demonstrated that plant lectins vary widely in their immunogenicity and in particular that mistletoe lectins (ML) I, II and II (MLI, MLII, MLIII) are potent immunogens when administered nasotracheally. Here, we measured immune responses following oral delivery of the MLs and assessed their ability to enhance responses to a co-administered antigen to determine if the molecules possess adjuvant activity. Oral administration of the lectins induced potent lectin-specific systemic and mucosal antibody responses. In addition, each of the three lectins possessed adjuvant activity when delivered orally together with ovalbumin (OVA). The lectins enhanced both serum and mucosal antibody responses to the co-delivered antigen. This shows for the first time that MLI, MLII and MLIII possess adjuvant activity when administered orally and may provide a platform for the generation of effective mucosal adjuvants.

The variability of TCM pattern diagnosis and herbal prescription on rheumatoid arthritis patients.

CONTEXT: The consistency of diagnosis made among Traditional Chinese Medicine (TCM) practitioners and the relationship between TCM diagnosis and Chinese herbal prescription have not been adequately examined. OBJECTIVE: To investigate the degree of consistency with which TCM diagnoses and herbal prescriptions can be made by practitioners examining rheumatoid arthritis (RA) patients. To survey TCM diagnostic patterns and to examine the correlation between herbal prescriptions and these diagnoses for a sample of RA patients. DESIGN: A prospective survey. SETTING: General Clinical Research Center, University of Maryland Hospital System, Baltimore, MD. PATIENTS: Rheumatoid arthritis patients. PRACTITIONERS: Licensed acupuncturists with a minimum of 5 years licensure and education in Chinese herbs. METHODS: Three TCM practitioners examined the same 39 RA patients separately, following the traditional "Four Diagnostic Methods." Patients filled out a questionnaire to serve as the data for the "Inquiry" component. They then underwent a physical examination, including the tongue and pulse, conducted by each of the practitioners. Based upon the examination results, each practitioner provided both a TCM diagnosis and a herbal prescription. These diagnoses/prescriptions were then examined with respect to the rate of agreement among the 3 practitioners. RESULTS: The average agreement with respect to the TCM diagnoses among the 3 pairs of TCM practitioners was 28.2% (25.6 to 33.3% with kappas ranging from 0.23 to 0.30). The degree to which the herbal prescriptions agreed with textbook recommended practice of each TCM diagnosis was 93.2% (range = 87.2 to 100%). CONCLUSION: The total agreement on TCM diagnosis on RA patients among 3 TCM practitioners was low. When less stringent, but theoretically justifiable, criteria were employed, greater consensus was obtained among the 3 practitioners. The correspondence between the TCM diagnosis and the herbal formula prescribed for that diagnosis was high, although there was little agreement among the 3 practitioners with respect to the herbal formulas prescribed for individual patients.

Analgesic duration and kinetics of liposomal bupivacaine after subcutaneous injection in mice.

1. The objective of the present study was to assess the time-course profile of analgesia and bupivacaine concentrations at the site of injection after subcutaneous administration of a single dose of standard bupivacaine or a novel controlled-release liposomal bupivacaine formulation. 2. Groups of mice were injected subcutaneously with 0.2 mL of 0.5% standard bupivacaine or 0.5, 1 or 2% liposomal bupivacaine. 3. A prolonged duration of analgesia occurred in mice receiving liposomal bupivacaine. In the liposomal groups, the bupivacaine remained at the injection site for more than 96 h, compared with approximately 8 h in groups injected with standard bupivacaine. 4. These results confirm that the prolonged analgesia observed after injection of the liposomal formulation is associated with sustained higher levels of bupivacaine at the site of injection.

Oral and intestinal mucositis - causes and possible treatments.

Chemotherapy and radiotherapy, whilst highly effective in the treatment of neoplasia, can also cause damage to healthy tissue. In particular, the alimentary tract may be badly affected. Severe inflammation, lesioning and ulceration can occur. Patients may experience intense pain, nausea and gastro-enteritis. They are also highly susceptible to infection. The disorder (mucositis) is a dose-limiting toxicity of therapy and affects around 500 000 patients world-wide annually. Oral and intestinal mucositis is multi-factorial in nature. The disruption or loss of rapidly dividing epithelial progenitor cells is a trigger for the onset of the disorder. However, the actual dysfunction that manifests and its severity and duration are greatly influenced by changes in other cell populations, immune responses and the effects of oral/gut flora. This complexity has hampered the development of effective palliative or preventative measures. Recent studies have concentrated on the use of bioactive/growth factors, hormones or interleukins to modify epithelial metabolism and reduce the susceptibility of the tract to mucositis. Some of these treatments appear to have considerable potential and are at present under clinical evaluation. This overview deals with the cellular changes and host responses that may lead to the development of mucositis of the oral cavity and gastrointestinal tract, and the potential of existing and novel palliative measures to limit or prevent the disorder. Presently available treatments do not prevent mucositis, but can limit its severity if used in combination. Poor oral health and existing epithelial damage predispose patients to mucositis. The elimination of dental problems or the minimization of existing damage to the alimentary tract, prior to the commencement of therapy, lowers their susceptibility. Measures that reduce the flora of the tract, before therapy, can also be helpful. Increased production of free radicals and the induction of inflammation are early events in the onset of mucositis. Prophylactic administration of scavengers or anti-inflammatories can partially counteract or limit some of these therapy-mediated effects, as can the use of cryotherapy. The regular use of mouthwashes, mouth coatings, antibiotics and analgesics is essential, prior to and during loss and ablation of the epithelial layer. Granulocyte-macrophage colony-stimulating factor/granulocyte colony-stimulating factor or the use of laser light therapy may aid restitution and repair. Glutamine supplements may be beneficial in the repair/recovery phase.

Assessing the consistency of traditional Chinese medical diagnosis: an integrative approach.

Traditional Chinese medicine (TCM) differs from its Western counterpart in a number of ways. One of the more striking ways involves a conceptually oriented diagnostic system that relies more on the clinician's reading of the patient's symptoms and signs than on laboratory findings. Because highly individualized TCM treatment plans emanate directly from its diagnostic system, a necessary condition for conducting clinically relevant TCM efficacy trials rests on the answer to a simple question: How consistent are different TCM practitioners at making the same TCM diagnoses and prescribing comparable treatment regimens for the same group of patients? Unfortunately, this question has not been adequately investigated, nor has the related question: Could the TCM diagnostic process be enhanced by access to modern biomedical tests? The purpose of this paper, therefore, is to discuss a number of the conceptual and methodological issues involved in the design of a recently funded NIH study whose primary purpose is to address these 2 questions.

Mistletoe lectins enhance immune responses to intranasally co-administered herpes simplex virus glycoprotein D2.

The mucosal adjuvant properties of the three type 2 ribosome-inactivating proteins (RIPs) from the European mistletoe, Viscum album L., were investigated. Mistletoe lectins were compared with cholera toxin (CT) as adjuvants when delivered nasotracheally together with herpes simplex virus glycoprotein D2 (gD2). All three mistletoe lectins (MLI, MLII, MLIII) were potent mucosal adjuvants. Co-administration of MLI, MLII or MLIII with gD2 led to significantly higher levels of gD2-specific mucosal immunoglobulin A (IgA) and systemic immunoglobulin G (IgG) antibody than when the antigen was delivered alone. The levels of antibodies induced were similar to those generated in mice immunized with gD2 and the potent mucosal adjuvant CT. Administration of ML1 with gD2 enhanced the antigen-specific splenic T-cell proliferative response. Interleukin-5 (IL-5), but not interferon-gamma (IFN-gamma), was detected in supernatants from splenocytes stimulated in vitro with gD2. This indicates that MLI enhanced type 2 T-helper cell (Th2) responses to the bystander antigen, gD2. Analysis of the gD2- and lectin-specific IgG subclass titres in mice immunized with gD2 and MLI, MLII or MLIII revealed a high ratio of IgG1 : IgG2a, which is compatible with the selective induction of Th2-type immune responses.