The Regulation of Aggrecanase ADAMTS-4 Expression in Human Achilles Tendon and tendon-Derived Cells

Several members of the ADAMTS (A Disintegrin And Metalloproteinase with ThromboSpondin motifs) family have been identified as aggrecanases, whose substrates include versican, the principal large proteoglycan in the tendon extracellular matrix. We have characterized the expression of ADAMTS-4 in human Achilles tendon and tendon-derived cells. ADAMTS-4 mRNA levels were higher in ruptured tendon compared with normal tendon or chronic painful tendinopathy. In tissue extracts probed by Western blotting, mature ADAMTS-4 (68 kDa) was detected only in ruptured tendons, while processed ADAMTS-4 (53 kDa) was detected also in chronic painful tendinopathy and in normal tendon. In cultured Achilles tendon cells, transforming growth factor-ß (TGF-ß) stimulated ADAMTS-4 mRNA expression (typically 20-fold after 24 h), while interleukin-1 induced a smaller, shorter-term stimulation which synergised markedly with that induced by TGF-ß. Increased levels of immunoreactive proteins consistent with mature and processed forms of ADAMTS-4 were detected in TGF-ß-stimulated cells. ADAMTS-4 mRNA was expressed at higher levels by tendon cells in collagen gels than in monolayer cultures. In contrast, the expression of ADAMTS-1 and -5 mRNA was lower in collagen gels compared with monolayers, and these mRNA showed smaller or opposite responses to growth factors and cytokines compared with that of ADAMTS-4 mRNA. We conclude that both ADAMTS-4 mRNA and ADAMTS-4 protein processing may be differentially regulated in normal and damaged tendons and that both the matrix environment and growth factors such as TGF-ß are potentially important factors controlling ADAMTS aggrecanase activities in tendon pathology

Inhibition of interleukin-1β-stimulated collagenase and stromelysin expression in human tendon fibroblasts by epigallocatechin gallate ester

The medicinal benefits of green tea (Camellia sinensis) consumption have been attributed to bioavailable polyphenols, notably epigallocatechin gallate (EGCG). We have assessed the effects of EGCG and its non-esterified counterpart EGC on the expression of the collagenases, matrix metalloproteinases (MMP)-1 and -13, and the stromelysin, MMP-3, in human tendon-derived fibroblasts. Interleukin (IL)-1ß increased MMP-1, -3 and -13 mRNA and output at least 30-fold. EGCG reduced this stimulation, by 20–30% at 2.5 µM and more than 80% at 25 µM, and had a smaller effect on MMP-2 mRNA expression, which was not stimulated by IL-1ß. In all experiments EGCG was at least 10-fold more potent than EGC. EGCG reduced the stimulation of p54 JNK/SAPK phosphorylation by IL-1ß but did not affect p38 MAPK phosphorylation, the degradation of I?B or the activating phosphorylation of NF?B. We conclude that EGCG reduces the IL-1-stimulated expression of both collagenase and stromelysin mRNA species, an effect which may be mediated by inhibition of the JNK/SAPK pathway. Taken together with previous reports of EGCG effects on the expression and/or activity of gelatinases and aggrecanases, our results underline the importance of extracellular matrix breakdown as a potential target for the actions of green tea polyphenols

Potent and Broad Inhibition of HIV-1 by a Peptide from the gp41 Heptad Repeat-2 Domain Conjugated to the CXCR4 Amino Terminus.

HIV-1 entry can be inhibited by soluble peptides from the gp41 heptad repeat-2 (HR2) domain that interfere with formation of the 6-helix bundle during fusion. Inhibition has also been seen when these peptides are conjugated to anchoring molecules and over-expressed on the cell surface. We hypothesized that potent anti-HIV activity could be achieved if a 34 amino acid peptide from HR2 (C34) were brought to the site of virus-cell interactions by conjugation to the amino termini of HIV-1 coreceptors CCR5 or CXCR4. C34-conjugated coreceptors were expressed on the surface of T cell lines and primary CD4 T cells, retained the ability to mediate chemotaxis in response to cognate chemokines, and were highly resistant to HIV-1 utilization for entry. Notably, C34-conjugated CCR5 and CXCR4 each exhibited potent and broad inhibition of HIV-1 isolates from diverse clades irrespective of tropism (i.e., each could inhibit R5, X4 and dual-tropic isolates). This inhibition was highly specific and dependent on positioning of the peptide, as HIV-1 infection was poorly inhibited when C34 was conjugated to the amino terminus of CD4. C34-conjugated coreceptors could also inhibit HIV-1 isolates that were resistant to the soluble HR2 peptide inhibitor, enfuvirtide. When introduced into primary cells, CD4 T cells expressing C34-conjugated coreceptors exhibited physiologic responses to T cell activation while inhibiting diverse HIV-1 isolates, and cells containing C34-conjugated CXCR4 expanded during HIV-1 infection in vitro and in a humanized mouse model. Notably, the C34-conjugated peptide exerted greater HIV-1 inhibition when conjugated to CXCR4 than to CCR5. Thus, antiviral effects of HR2 peptides can be specifically directed to the site of viral entry where they provide potent and broad inhibition of HIV-1. This approach to engineer HIV-1 resistance in functional CD4 T cells may provide a novel cell-based therapeutic for controlling HIV infection in humans

Expression profiling of metalloproteinases and tissue inhibitors of metalloproteinases in normal and degenerate human achilles tendon

To profile the messenger RNA (mRNA) expression for the 23 known genes of matrix metalloproteinases (MMPs), 19 genes of ADAMTS, 4 genes of tissue inhibitors of metalloproteinases (TIMPs), and ADAM genes 8, 10, 12, and 17 in normal, painful, and ruptured Achilles tendons. Tendon samples were obtained from cadavers or from patients undergoing surgical procedures to treat chronic painful tendinopathy or ruptured tendon. Total RNA was extracted and mRNA expression was analyzed by quantitative real-time reverse transcription–polymerase chain reaction, normalized to 18S ribosomal RNA. In comparing expression of all genes, the normal, painful, and ruptured Achilles tendon groups each had a distinct mRNA expression signature. Three mRNA were not detected and 14 showed no significant difference in expression levels between the groups. Statistically significant (P < 0.05) differences in mRNA expression, when adjusted for age, included lower levels of MMPs 3 and 10 and TIMP-3 and higher levels of ADAM-12 and MMP-23 in painful compared with normal tendons, and lower levels of MMPs 3 and 7 and TIMPs 2, 3, and 4 and higher levels of ADAMs 8 and 12, MMPs 1, 9, 19, and 25, and TIMP-1 in ruptured compared with normal tendons. The distinct mRNA profile of each tendon group suggests differences in extracellular proteolytic activity, which would affect the production and remodeling of the tendon extracellular matrix. Some proteolytic activities are implicated in the maintenance of normal tendon, while chronically painful tendons and ruptured tendons are shown to be distinct groups. These data will provide a foundation for further study of the role and activity of many of these enzymes that underlie the pathologic processes in the tendon

Relationships between Irritable Bowel Syndrome Pain, Skin Temperature Indices of Autonomic Dysregulation, and Sensitivity to Thermal Cutaneous Stimulation

This study evaluated relationships between irritable bowel syndrome (IBS) pain, sympathetic dysregulation, and thermal pain sensitivity. Eight female patients with diarrhea-predominant IBS and ten healthy female controls were tested for sensitivity to thermal stimulation of the left palm. A new method of response-dependent thermal stimulation was used to maintain pain intensity at a predetermined level (35%) by adjusting thermal stimulus intensity as a function of pain ratings. Clinical pain levels were assessed prior to each testing session. Skin temperatures were recorded before and after pain sensitivity testing. The temperature of palmar skin dropped (1.5°C) when the corresponding location on the opposite hand of control subjects was subjected to prolonged thermal stimulation, but this response was absent for IBS pain patients. The patients also required significantly lower stimulus temperatures than controls to maintain a 35% pain rating. Baseline skin temperatures of patients were significantly correlated with thermode temperatures required to maintain 35% pain ratings. IBS pain intensity was not significantly correlated with skin temperature or pain sensitivity. The method of response-dependent stimulation revealed thermal hyperalgesia and increased sympathetic tone for chronic pain patients, relative to controls. Similarly, a significant correlation between resting skin temperatures and thermal pain sensitivity for IBS but not control subjects indicates that tonic sympathetic activation and a thermal hyperalgesia were generated by the chronic presence of visceral pain. However, lack of a significant relationship between sympathetic tone and ratings of IBS pain casts doubt on propositions that the magnitude of IBS pain is determined by psychological stress

Criteria for the diagnosis of corticobasal degeneration

Current criteria for the clinical diagnosis of pathologically confirmed corticobasal degeneration (CBD) no longer reflect the expanding understanding of this disease and its clinicopathologic correlations. An international consortium of behavioral neurology, neuropsychology, and movement disorders specialists developed new criteria based on consensus and a systematic literature review. Clinical diagnoses (early or late) were identified for 267 nonoverlapping pathologically confirmed CBD cases from published reports and brain banks. Combined with consensus, 4 CBD phenotypes emerged: corticobasal syndrome (CBS), frontal behavioral-spatial syndrome (FBS), nonfluent/agrammatic variant of primary progressive aphasia (naPPA), and progressive supranuclear palsy syndrome (PSPS). Clinical features of CBD cases were extracted from descriptions of 209 brain bank and published patients, providing a comprehensive description of CBD and correcting common misconceptions. Clinical CBD phenotypes and features were combined to create 2 sets of criteria: more specific clinical research criteria for probable CBD and broader criteria for possible CBD that are more inclusive but have a higher chance to detect other tau-based pathologies. Probable CBD criteria require insidious onset and gradual progression for at least 1 year, age at onset ≥50 years, no similar family history or known tau mutations, and a clinical phenotype of probable CBS or either FBS or naPPA with at least 1 CBS feature. The possible CBD category uses similar criteria but has no restrictions on age or family history, allows tau mutations, permits less rigorous phenotype fulfillment, and includes a PSPS phenotype. Future validation and refinement of the proposed criteria are needed

Longitudinal Bioluminescence Imaging of Primary Versus Abdominal Metastatic Tumor Growth in Orthotopic Pancreatic Tumor Models in NSG Mice

Objectives: The purpose of the present study was to develop and validate noninvasive bioluminescence imaging methods for differentially monitoring primary and abdominal metastatic tumor growth in mouse orthotopic models of pancreatic cancer. Methods: A semiautomated maximum entropy segmentation method was implemented for the primary tumor region of interest, and a rule-based method for manually drawing a region of interest for the abdominal metastatic region was developed for monitoring tumor growth in orthotopic models of pancreatic cancer. The 2 region-of-interest methods were validated by having 2 observers independently segment Panc-1 tumors, and the results were compared with the number of mesenteric lymph node nodules and histopathologic assessment of liver metastases. The findings were extended to orthotopic tumors of the more metastatic MIA PaCa-2 and AsPC-1 cells where separate groups of animals were implanted with different numbers of cells. Results: The results demonstrated that the segmentation methods were highly reliable, reproducible, and robust and allowed statistically significant discrimination in the growth rates of primary and abdominal metastatic tumors of different cell lines implanted with different numbers of cells. Conclusions: The present results demonstrate that primary tumors and abdominal metastatic foci in orthotopic pancreatic cancer models can be reliably quantified separately and noninvasively over time with bioluminescence imaging

Layperson perception of reflux-related symptoms

OBJECTIVE: To assess for differences of intended meaning in the description of reflux-related symptoms among otolaryngology patients and clinicians. STUDY DESIGN: Cross-sectional survey-based study. SETTING: Five tertiary, academic otolaryngology practices. METHODS: Between June 2020 and July 2022, a questionnaire consisting of 20 common descriptors of reflux-related symptoms within four domains (throat-, chest-, stomach-, and sensory-related symptoms) was completed by patients. Attending otolaryngologists at five academic medical centers then completed the same survey. The primary outcome was to assess differences in patient and clinician perceptions of reflux-related symptoms. Differences based on geographic location was a secondary outcome. RESULTS: A total of 324 patients and 27 otolaryngologists participated. Patients selected a median of six terms compared with 10.5 for otolaryngologists ( CONCLUSION: There are differences between otolaryngologists and their patients in the interpretation of the symptoms of reflux. Patients tended to have a narrower interpretation of reflux with symptoms primarily limited to classic stomach-related symptoms, while clinicians tended to have a broader definition of reflux that included extra-esophageal manifestations of disease. This has important counseling implications for the clinician, as patients presenting with reflux symptoms may not comprehend the relationship of those symptoms to reflux disease

Interactions of Bacillus Mojavensis and Fusarium Verticillioides With a Benzoxazolinone (Boa) and Its Transformation Product, Apo

En:Journal of Chemical Ecology (2007, vol. 33, n. 10, p. 1885-1897)The benzoxazolinones, specifically benzoxazolin-2(3H)-one (BOA), are important transformation products of the benzoxazinones that can serve as allelochemicals providing resistance to maize from pathogenic bacteria, fungi, and insects. However, maize pathogens such as Fusarium verticillioides are capable of detoxifying the benzoxazolinones to 2-aminophenol (AP), which is converted to the less toxic N-(2-hydroxyphenyl) malonamic acid (HPMA) and 2-acetamidophenol (HPAA). As biocontrol strategies that utilize a species of endophytic bacterium, Bacillus mojavensis, are considered efficacious as a control of this Fusarium species, the in vitro transformation and effects of BOA on growth of this bacterium was examined relative to its interaction with strains of F. verticillioides. The results showed that a red pigment was produced and accumulated only on BOA-amended media when wild type and the progeny of genetic crosses of F. verticillioides are cultured in the presence of the bacterium. The pigment was identified as 2-amino-3H-phenoxazin-3-one (APO), which is a stable product. The results indicate that the bacterium interacts with the fungus preventing the usual transformation of AP to the nontoxic HPMA, resulting in the accumulation of higher amounts of APO than when the fungus is cultured alone. APO is highly toxic to F. verticillioides and other organisms. Thus, an enhanced biocontrol is suggested by this in vitro study. =580 $aEn:Journal of Chemical Ecolog

An Arthroscopic Device to Assess Articular Cartilage Defects and Treatment with a Hydrogel

The hydraulic resistance R across osteochondral tissue, especially articular cartilage, decreases with degeneration and erosion. Clinically useful measures to quantify and diagnose the extent of cartilage degeneration and efficacy of repair strategies, especially with regard to pressure maintenance, are still developing. The hypothesis of this study was that hydraulic resistance provides a quantitative measure of osteochondral tissue that could be used to evaluate the state of cartilage damage and repair. The aims were to (1) develop a device to measure R in an arthroscopic setting, (2) determine whether the device could detect differences in R for cartilage, an osteochondral defect, and cartilage treated using a hydrogel ex vivo, and (3) determine how quickly such differences could be discerned. The apparent hydraulic resistance of defect samples was ~35% less than intact cartilage controls, while the resistance of hydrogel-filled groups was not statistically different than controls, suggesting some restoration of fluid pressurization in the defect region by the hydrogel. Differences in hydraulic resistance between control and defect groups were apparent after 4 s. The results indicate that the measurement of R is feasible for rapid and quantitative functional assessment of the extent of osteochondral defects and repair. The arthroscopic compatibility of the device demonstrates the potential for this measurement to be made in a clinical setting