Search

The aim of the circular economy is to treat waste as a valuable raw material, reintegrating it into the industrial economy and extending the lifecycle of subsequent products. Efforts to reduce the production of hard-to-recycle waste are becoming increasingly important to manufacturers, not only of consumer goods but also of specialized items that are difficult to manufacture, such as medical supplies, which have now become a priority for the European Union. The purpose of the study is to manufacture a novel human-purified type I collagen membrane from bone remnants typically discarded during the processing of cortico-cancellous bones in tissue banks and to evaluate its mechanical properties and effectiveness in regenerating bone-critical mandibular defects in rabbits. To prepare the novel membrane, cortico-cancellous bone chip samples from a local tissue bank were processed to isolate collagen by demineralization under agitation in HCl, cast into a silicone mold, and air-dried at room temperature and UV irradiation. The average thickness of the four batches analyzed by SEM was 37.3 μm. The average value of Young's modulus and tensile strength obtained from the specimens was 2.56 GPa and 65.43 Mpa, respectively. The membrane's efficacy was tested by creating a critical bicortical and bilateral osteoperiosteal defect in rabbit mandibles. The right-side defects were covered with the collagen membrane, while the left-side defects were left untreated as a control. Nine weeks post-surgery, clinical, radiological, and histological analyses demonstrated new bone formation in the treated areas, whereas the control sites showed no bone regeneration. This innovative approach not only contributes to sustainability in healthcare by optimizing biological waste but also exemplifies efficient resource use in line with the circular economy, offering a cost-effective, biocompatible option that could benefit national health systems.

Introduction: In-stent restenosis (ISR) is a major challenge in interventional cardiology. Both ISR and excessive skin healing are aberrant hyperplasic responses, which may be functionally related. However, the cellular component underlying ISR remains unclear, especially regarding vascular homeostasis. Recent evidence suggest that novel immune cell populations may be involved in vascular repair and damage, but their role in ISR has not been explored. The aims of this study is to analyze (i) the association between ISR and skin healing outcomes, and (ii) the alterations in vascular homeostasis mediators in ISR in univariate and integrative analyses., Methods: 30 patients with ≥1 previous stent implantation with restenosis and 30 patients with ≥1 stent without restenosis both confirmed in a second angiogram were recruited. Cellular mediators were quantified in peripheral blood by flow cytometry. Skin healing outcomes were analyzed after two consecutive biopsies., Results: Hypertrophic skin healing was more frequent in ISR patients (36.7%) compared to those ISR-free (16.7%). Patients with ISR were more likely to develop hypertrophic skin healing patterns (OR 4.334 [95% CI 1.044-18.073], p=0.033), even after correcting for confounders. ISR was associated with decreased circulating angiogenic T-cells (p=0.005) and endothelial progenitor cells (p<0.001), whereas CD4 + CD28 null and detached endothelial cells counts were higher (p<0.0001 and p=0.006, respectively) compared to their ISR-free counterparts. No differences in the frequency of monocyte subsets were found, although Angiotensin-Converting Enzyme expression was increased (non-classical: p<0.001; and intermediate: p<0.0001) in ISR. Despite no differences were noted in Low-Density Granulocytes, a relative increase in the CD16 - compartment was observed in ISR (p=0.004). An unsupervised cluster analysis revealed the presence of three profiles with different clinical severity, unrelated to stent types or traditional risk factors., Conclusion: ISR is linked to excessive skin healing and profound alterations in cellular populations related to vascular repair and endothelial damage. Distinct cellular profiles can be distinguished within ISR, suggesting that different alterations may uncover different ISR clinical phenotypes., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Lozano, Bangueses, Rodríguez, Pevida, Rodríguez-Aguilar, Rodríguez, Espasandín-Arias, Llames, Meana, Suárez and Rodríguez-Carrio.)

Background: Cancer-associated fibroblasts (CAFs) are major players in tumor-stroma communication, and participate in several cancer hallmarks to drive tumor progression and metastatic dissemination. This study investigates the driving effects of tumor-secreted factors on CAF biology, with the ultimate goal of identifying effective therapeutic targets/strategies for head and neck squamous cell carcinomas (HNSCC)., Methods: Functionally, conditioned media (CM) from different HNSCC-derived cell lines and normal keratinocytes (Kc) were tested on the growth and invasion of populations of primary CAFs and normal fibroblasts (NFs) using 3D invasion assays in collagen matrices. The changes in MMPs expression were evaluated by RT-qPCR and kinase enrichment was analyzed using mass spectrometry phosphoproteomics., Results: Our results consistently demonstrate that HNSCC-secreted factors (but not Kc CM) specifically and robustly promoted pro-invasive properties in both CAFs and NFs, thereby reflecting the plasticity of fibroblast subtypes. Concomitantly, HNSCC-secreted factors massively increased metalloproteinases levels in CAFs and NFs. By contrast, HNSCC CM and Kc CM exhibited comparable growth-promoting effects on stromal fibroblasts. Mechanistically, phosphoproteomic analysis predominantly revealed phosphorylation changes in fibroblasts upon treatment with HNSCC CM, and various promising kinases were identified: MKK7, MKK4, ASK1, RAF1, BRAF, ARAF, COT, PDK1, RSK2 and AKT1. Interestingly, pharmacologic inhibition of RAF1/BRAF using sorafenib emerged as the most effective drug to block tumor-promoted fibroblast invasion without affecting fibroblast viability CONCLUSIONS: Our findings demonstrate that HNSCC-secreted factors specifically fine tune the invasive potential of stromal fibroblasts, thereby generating tumor-driven pro-invasive niches, which in turn to ultimately facilitate cancer cell dissemination. Furthermore, the RAF/BRAF inhibitor sorafenib was identified as a promising candidate to effectively target the onset of pro-invasive clusters of stromal fibroblasts in the HNSCC microenvironment., Competing Interests: Conflict of Interest Statement The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

The rabbit skin irritation test has been the standard for evaluating the irritation potential of chemicals; however, alternative methods that do not use animal testing are actively encouraged. Reconstructed human epidermis (RhE) models mimic the biochemical and physiological properties of the human epidermis and can be used as an alternative method. On RhE methods, the metabolic activity of RhE models is used to predict skin irritation, with a reduction in metabolic activity indicating a reduced number of viable cells and linking cell death to skin irritation processes. However, new challenges have emerged as the use of RhE models increases, including the need for non-invasive and marker-free methodologies to assess cellular states. Electrochemical impedance spectroscopy (EIS) is one such methodology that can meet these requirements. In this study, our results showed that EIS can differentiate between irritant and non-irritant chemicals, with a significant increase in the capacitance values observed in the irritant samples. A ROC curve analysis showed that the prediction method based on EIS met OECD TG 439 requirements at all time points and had 95% within-laboratory reproducibility. Comparison with the MTT viability assay showed that prediction using EIS achieved higher sensitivity, specificity, and accuracy. These results suggest that EIS could potentially replace animal testing in the evaluation of irritation potential and could be a valuable addition to in vitro testing strategies.

Reconstructive surgery techniques have evolved exponentially in last decades. From regional flaps to free tissue transfer, tissue movilization has become the gold standart treatment in many reconstructive procedures. Main disadvantage from these techniques lies in the possibility of sequels in donor zone. Furthermore, raising comorbidities in general population and growing indications for reconstructive surgery in elder people, have triggered the development of new biomaterials which can offer support in the reconstruction while elicit donor zone morbidity. Advances in tissue decellularization techniques have brought numerous matrices which have shown effectivity in many reconstructive procedures. Use of acellular dermal matrices may become an eligible solution for many reconstructive procedures. From breast reconstruction assisted by matrices to complex wound coverage passing throught tendon repair techniques, acellular dermal matrices have shown effectiveness in last studies. Local production of this biomaterial leads to cost minimization derived from harvesting and manufacturing matrices in our centre and avoid out-of-stock and storage issues. Current original protocol proposed by our group include all steps from harvesting samples from cadaveric donors till matrix storage after decellularization proccess. The result is a high valued biomaterial in terms of biocompatibility and security profile available., (© 2021. The Author(s), under exclusive licence to Springer Nature B.V.)

The skin is the largest organ in the human body, comprising the main barrier against the environment. When the skin loses its integrity, it is critical to replace it to prevent water loss and the proliferation of opportunistic infections. For more than 40 years, tissue-engineered skin grafts have been based on the in vitro culture of keratinocytes over different scaffolds, requiring between 3 to 4 weeks of tissue culture before being used clinically. In this study, we describe the development of a polymerizable skin hydrogel consisting of keratinocytes and fibroblast entrapped within a fibrin scaffold. We histologically characterized the construct and evaluated its use on an in vivo wound healing model of skin damage. Our results indicate that the proposed methodology can be used to effectively regenerate skin wounds, avoiding the secondary in vitro culture steps and thus, shortening the time needed until transplantation in comparison with other bilayer skin models. This is achievable due to the instant polymerization of the keratinocytes and fibroblast combination that allows a direct application on the wound. We suggest that the polymerizable skin hydrogel is an inexpensive, easy and rapid treatment that could be transferred into clinical practice in order to improve the treatment of skin wounds.

Background: Cell surface glycosaminoglycans (GAGs) participate in many physiological and pathological processes, including infections and inflammatory response. Acne is a common chronic inflammatory skin disorder that affects the pilosebaceous unit and has a multifactorial etiology, including bacterial colonization of the hair follicle. This study aimed to investigate the participation of GAG in the adhesion of Propionibacterium acnes, Staphylococcus aureus and Staphylococcus epidermidis to keratinocytes and fibroblasts of the skin by competition experiments and cell surface removal using specific liases. The alteration in the transcription of the genes responsible for the synthesis of GAG induced by the adhesion of these bacteria was also analyzed by qRT-PCR., Results: GAGs are involved in bacterial adherence to skin cells, especially fibroblasts, where chondroitin sulfate displayed the higher effect. Bacterial adherence produced different alterations in the transcription of the genes responsible for GAG structures. P. acnes induced mostly changes in keratinocytes, while S. epidermidis was the main cause of alterations in fibroblasts. These variations in gene expression affected all the stages in the biosynthesis of the main species of GAGs, heparan and chondroitin sulphate., Conclusions: GAGs species are involved in the adhesion of acne-related bacteria to skin cells in a differential manner depending on each microorganism and cellular type, although other receptors seem to exist. Bacterial adherence led to variations on gene expression in skin cells affecting GAG chains structure what, consequently, should alter their interactions with different ligands, affecting the development of acne disease., (© 2022. The Author(s).)

Familial melanoma accounts for 10% of cases, being CDKN2A the main high-risk gene. However, the mechanisms underlying melanomagenesis in these cases remain poorly understood. Our aim was to analyze the transcriptome of melanocyte-keratinocyte co-cultures derived from healthy skin from familial melanoma patients vs. controls, to unveil pathways involved in melanoma development in at-risk individuals. Accordingly, primary melanocyte-keratinocyte co-cultures were established from the healthy skin biopsies of 16 unrelated familial melanoma patients (8 CDKN2A mutant, 8 CDKN2A wild-type) and 7 healthy controls. Whole transcriptome was captured using the SurePrint G3 Human Microarray. Transcriptome analyses included: differential gene expression, functional enrichment, and protein-protein interaction (PPI) networks. We identified a gene profile associated with familial melanoma independently of CDKN2A germline status. Functional enrichment analysis of this profile showed a downregulation of pathways related to DNA repair and immune response in familial melanoma ( P < 0.05). In addition, the PPI network analysis revealed a network that consisted of double-stranded DNA repair genes (including BRCA1, BRCA2, BRIP1 , and FANCA ), immune response genes, and regulation of chromosome segregation. The hub gene was BRCA1 . In conclusion, the constitutive deregulation of BRCA1 pathway genes and the immune response in healthy skin could be a mechanism related to melanoma risk., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Potrony, Haddad, Tell-Martí, Gimenez-Xavier, Leon, Pevida, Mateu, Badenas, Carrera, Malvehy, Aguilera, Llames, Escámez, Puig-Butillé, del Río and Puig.)

Defining the corrosive properties of chemical products generally involves the use of animal models for human health safety assessment. However, a few alternatives to animal experimentation are currently internationally accepted in order to reduce animal suffering. One of these alternatives makes use of in vitro reconstructed human epidermis (RhE) models and predicts corrosive potential based on the evaluation of cell viability after topical exposure. These models rely on its similarity to human skin, both functional and histological, and are currently worldwide marketed by a few private companies. In this manuscript, we describe the fundamentals of the production of a Do It Yourself (DIY) RhE model, and the operating procedures for the assessment of skin corrosion based on the guidelines proposed for the development of new alternative methods for skin corrosion. Our results indicate that the DIY-RhE model resembles the anatomy of the normal human epidermis as seen by immunohistochemical analysis. Moreover, barrier properties of DIY-RhE were assessed by the measure of Transepithelial Electrical Resistance. Applicability of DIY-RhE for the assessment of skin corrosion was evaluated by measuring cell viability after topical exposure of twelve reference chemicals for 3 and 60 min. Predictive performance resulted in 100% sensitivity, 100% specificity and 100% accuracy matching current requirements for new RhE models proposed for the discrimination of corrosives and non-corrosives., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Tissue engineering is one of the fields of clinical medicine that has forged ahead in recent years, especially because of its role as a potential alternative to organ transplantation. The main aim of this study has been the development of biocompatible materials to form extracellular matrix (ECM) structures in order to provide the necessary conditions for the settlement, proliferation and differentiation of dermal cells such as fibroblasts. To this end, human plasma gels were synthesized with the addition of increasing concentrations of transglutaminase (TGase), which catalyses the formation of covalent bonds between Lys and Glu residues. These materials were structurally characterized using rheology and texturometry and were found to have good structural resistance and elasticity for fibroblast culture. A remarkable improvement in the mechanical properties of the human plasma gels was detected when the two highest TGase concentrations were tested, which may be interpreted as an increase in the number of covalent and non-covalent bonds formed between the plasma protein chains. Furthermore, a human fibroblast primary culture was seeded on human plasma scaffolds and satisfactorily proliferated at 37 °C. This was verified in the images obtained by optical microscopy (OM) and by scanning electron microscopy (SEM), which confirmed that the structure of this type of material is suitable for the growth and proliferation of dermal fibroblasts., (Copyright © 2018. Published by Elsevier Ltd.)

Chemokine CCL2, also known as monocyte chemoattractant protein-1 (MCP-1), is a molecule that in addition to its well-established role in chemotaxis can also act as nociceptor sensitizer. The upregulation of this chemokine in inflamed tissues could suggest its involvement in inflammatory hypernociception. Thus, we have measured CCL2 levels in mice with acute or chronic inflammation due to the intraplantar (i.pl.) injection of carrageenan or complete Freund's adjuvant (CFA), respectively, and we have studied whether inflammatory hyperalgesia or allodynia could be attenuated by blocking CCR2 receptors or neutralizing CCL2 with an anti-CCL2 antibody. A remarkable increase in CCL2 concentration was detected by ELISA in paw homogenates coming from carrageenan- or CFA-inflamed mice, being its expression mainly localized in macrophages, as shown by immunohistochemical assays. The s.c. (0.3-3 mg/kg) or i.pl. (0.3-3 μg) administration of the CCR2 antagonist, RS 504393, dose dependently inhibited thermal hyperalgesia measured in acutely or chronically inflamed mice, whereas s.c. administration of this drug did not reduce inflammatory mechanical allodynia. Furthermore, the inhibition of inflammatory hyperalgesia after the administration of an anti-CCL2 antibody (0.1-1 μg; i.pl.) suggests that CCL2 could be the endogenous chemokine responsible for CCR2-mediated hyperalgesic effects. Besides, the acute administration of the highest antihyperalgesic dose of RS 504393 assayed did not reduce paw tumefaction or modify the presence of inflammatory cells. These results indicate that the blockade of the CCL2/CCR2 system can counteract inflammatory hyperalgesia, being this antinociceptive effect unrelated to a decrease in the inflammatory reaction., (© 2016 Société Française de Pharmacologie et de Thérapeutique.)

Background: Pain due to bone metastases of prostatic origin is a relevant clinical issue. We study here the nociceptive responses obtained in mice receiving the intratibial inoculation of RM1 prostate cancer cells., Methods: 10(2) -10(5) RM1 cells were inoculated to C57BL/6 mice and tumor development was analysed histologically and with luciferase-expressing RM1 cells. Spinal astroglial (GFAP) or microglial (Iba-1) expression was assessed with immunohistochemical methods and hypernociception was measured by the unilateral hot plate, the paw pressure and the von Frey tests. The analgesic effect of morphine, zoledronic acid or the CCR2 antagonist RS504393 was measured. Levels of the chemokines CCL2, CCL3, and CCL5 were determined by ELISA., Results: The inoculation of 10(3) RM1 cells induced tumoral growth in bone with a mixed osteoclastic/osteoblastic pattern and evoked astroglial, but not microglial, activation in the spinal cord. Hyperalgesia and allodynia were already established four days after inoculation and dose-dependently inhibited by the s.c. administration of morphine (1-5 mg/kg) or zoledronic acid (1-3 mg/kg). CCL2 and CCL5, but not CCL3, were released by RM1 cells in culture whereas only an increased presence of CCL2 was found in bone tumor homogenates. The administration of the CCR2 antagonist RS504393 (0.3-3 mg/kg) inhibited RM1 induced thermal hyperalgesia without modifying mechanical allodynia., Conclusion: The intratibial inoculation of RM1 cells in immunocompetent mice induces hypernociceptive responses and can be useful to perform studies of bone cancer induced pain related to androgen-independent prostate cancer. The antinociceptive role derived from the blockade of the CCR2 chemokine receptors is further envisaged., (© 2014 Wiley Periodicals, Inc.)

Germline mutations in CDKN2A and/or red hair color variants in MC1R genes are associated with an increased susceptibility to develop cutaneous melanoma or non melanoma skin cancer. We studied the impact of the CDKN2A germinal mutation p.G101W and MC1R variants on gene expression and transcription profiles associated with skin cancer. To this end we set-up primary skin cell co-cultures from siblings of melanoma prone-families that were later analyzed using the expression array approach. As a result, we found that 1535 transcripts were deregulated in CDKN2A mutated cells, with over-expression of immunity-related genes (HLA-DPB1, CLEC2B, IFI44, IFI44L, IFI27, IFIT1, IFIT2, SP110 and IFNK) and down-regulation of genes playing a role in the Notch signaling pathway. 3570 transcripts were deregulated in MC1R variant carriers. In particular, genes related to oxidative stress and DNA damage pathways were up-regulated as well as genes associated with neurodegenerative diseases such as Parkinson's, Alzheimer and Huntington. Finally, we observed that the expression signatures indentified in phenotypically normal cells carrying CDKN2A mutations or MC1R variants are maintained in skin cancer tumors (melanoma and squamous cell carcinoma). These results indicate that transcriptome deregulation represents an early event critical for skin cancer development.

The hypernociceptive role played by the chemokine CCL2, and its main receptor, CCR2, in pathological settings is being increasingly recognized. We aimed to characterize the involvement of spinal CCL2 in the hyperalgesia due to the intratibial inoculation of fibrosarcoma NCTC 2472 cells in mice. The intrathecal (i.t.) administration of the CCR2 antagonist RS 504393 (1–3 μg) or an anti-CCL2 antibody inhibited tumoral hyperalgesia. No change in the expression of spinal CCR2 was detected by western blot, whereas immunohistochemical experiments demonstrated increased CCL2 staining at the superficial laminae of the spinal cord ipsilateral to the tumor. This spinal CCL2 does not seem to be released from nociceptors since CCL2 mRNA and CCL2 levels in DRGs, as measured by RT-PCR and ELISA, remain unmodified in tumor-bearing mice. In contrast, immunohistochemical assays demonstrated the spinal up-regulations of GFAP and Iba-1, respective markers of astroglia and microglia, and the expression of CCL2 in both types of glial cells at the superficial laminae of the spinal cord of tumor-bearing mice. Finally, since CCL2 could induce astroglial or microglial activation, we studied whether the blockade of CCR2 could inhibit the increased spinal glial expression. GFAP, but not Iba-1, up-regulation was reduced in tumor-bearing mice treated for 3 days with i.t. RS 504393, indicating that spinal CCL2 acts as an astroglial activator in this setting. The participation at spinal level of CCL2/CCR2 in tumoral hypernociception, together with its previously described involvement at periphery, makes attractive the modulation of this system for the alleviation of neoplastic pain.

The antineoplastic paclitaxel induces a sensory neuropathy that involves the spinal release of neuroinflammatory mediators and activation of glial cells. Although the chemokine CCL2 can evoke glial activation and its participation in neuropathic pain has been demonstrated in other models, its involvement in paclitaxel-evoked neuropathy has not been previously explored. Paclitaxel-evoked cold hypernociception was assessed in mice by the unilateral cold plate test and the effects on cold hyperalgesia of the CCR2 antagonist RS 504393, the CCR1 antagonist J113863, the microglial inhibitor minocycline or an anti-CCL2 antibody were tested. Furthermore, ELISA measurements of CCL2 concentration and immunohistochemical assays of Iba-1 and GFAP, markers of microglial and astroglial cells respectively, were performed in the lumbar spinal cord. Cold hypernociception measured 3 days after the administration of paclitaxel (10mg/kg) was inhibited by the s.c. (0.3-3mg/kg) or i.t. (1-10 μg) administration of RS 504393 but not of J113863 (3-30 mg/kg). CCL2 levels measured by ELISA in the lumbar spinal cord were augmented in mice treated with paclitaxel and the i.t. administration of an anti-CCL2 antibody completely suppressed paclitaxel-evoked cold hyperalgesia, strongly suggesting that CCL2 is involved in the hypernociception evoked by this taxane. Besides, the implication of microglial activation is supported by the increase in the immunolabelling of Iba-1, but not GFAP, in the spinal cord of paclitaxel-treated mice and by the inhibition of cold hyperalgesia produced by the i.t. administration of the microglial inhibitor minocycline (1-10 nmol). Finally, the neutralization of spinal CCL2 by the i.t. administration of a selective antibody for 3 days almost totally inhibited paclitaxel-evoked microglial activation. In conclusion, our results indicate that paclitaxel-evoked cold hypernociception depends on the activation of CCR2 due to the spinal release of CCL2 and the subsequent microglial activation., (Copyright © 2013 Elsevier Inc. All rights reserved.)

The participation of the chemokine CCL2 (monocyte chemoattractant protein-1) in inflammatory and neuropathic pain is well established. Furthermore, the release of CCL2 from a NCTC 2472 cells-evoked tumor and its involvement in the upregulation of calcium channel α2δ1 subunit of nociceptors was demonstrated. In the present experiments, we have tried to determine whether the increase in CCL2 levels is a common property of painful tumors and, in consequence, the administration of a chemokine receptor type 2 (CCR2) antagonist can inhibit tumoral hypernociception. CCL2 levels were measured by ELISA in the tumoral region of mice intratibially inoculated with NCTC 2472 or B16-F10 cells, and the antihyperalgesic and antiallodynic effects evoked by the administration of the selective CCR2 antagonist RS 504393 were assessed. Cultured NCTC 2472 cells release CCL2 and their intratibial inoculation evokes the development of a tumor in which CCL2 levels are increased. Moreover, the systemic or peritumoral administration of RS 504393 inhibited thermal and mechanical hyperalgesia, but not mechanical allodynia evoked after the inoculation of these cells. Thermal hyperalgesia was also inhibited by the peritumoral administration of a neutralizing CCL2 antibody. In contrast, no change in CCL2 levels was observed in mice inoculated with B16-F10 cells, and RS 504393 did not inhibit the hypernociceptive reactions evoked by their intratibial inoculation. The peripheral release of CCL2 is involved in the development of thermal and mechanical hyperalgesia, but not mechanical allodynia evoked by the inoculation of NCTC 2472 cells, whereas this chemokine seems unrelated to the hypernociception induced by B16-F10 cells.

Osteomyelitis (OM) is a bone infection characterized by necrosis and new formation of bone. Because matrix metalloproteases (MMPs) play an important role in bone extracellular matrix remodeling, we investigated the role of some MMP polymorphisms in OM patients. A total of 118 OM patients and 300 blood donors were genotyped for the polymorphisms of MMP1 (-1607 1G/2G) and MMP13 (-77A/G). Levels of MMPs (-1, -2, -3, -8, -9, -10, and -13) and tissue inhibitors of metaloproteases (TIMP-1, -2, and -4) in serum and in human osteoblasts obtained from OM biopsies also were determined. The MMP1 (-1607 2G/2G) genotype was significantly more frequent among OM patients compared with controls [65.3% versus 33.7%, chi(2) = 26.85, odds ratio (OR) = 3.24, 95% confidence interval (CI) 2.03-5.2, p < .0001]. The MMP1 2G allele also was more frequent in OM patients (73.3% versus 57.2%, chi(2) = 37.76, OR = 2.75, 95% CI 1.96-3.85, p < .0001). Carriers of the 2G allele had significantly higher osteoblast MMP1 mRNA and MMP-1 serum levels than noncarriers (p < .04). Interleukin 1alpha (IL-1alpha) increased MMP-1 and -13 protein secretion and Ets1 mRNA expression by OM patients' osteoblasts. No association of the MMP13 (-77 A/G) polymorphism with OM was observed. The MMP1 (-1607 1G/2G) polymorphism might contribute to OM pathogenesis. This could be due to increased expression of MMP-1 by osteoblasts and is regulated by IL-1alpha., (Copyright 2010 American Society for Bone and Mineral Research.)

Although previous studies describe the up-regulation of purinergic P2X(3) receptors expressed at peripheral nociceptive fibers in experimental painful neoplastic processes, the analgesic efficacy of P2X(3) receptor antagonists has not been tested in these settings. We study here the effect of the P2X(3) receptor antagonist, A-317491, on thermal hyperalgesia produced by the intratibial inoculation of NCTC 2472 fibrosarcoma cells to C3H/HeJ mice. The peritumoral administration of A-317491 (10-100 microg) dose-dependently attenuated osteosarcoma-induced thermal hyperalgesia without modifying thermal latencies measured in the contralateral paws. This antihyperalgesic effect was inhibited by the coadministration of naloxone-methiodide (0.1-1 microg) or the systemic injection of the selective mu-opioid receptor antagonist cyprodime (1 mg/kg), demonstrating the involvement of peripheral mu-opioid receptors. Furthermore, the antihyperalgesic effect induced by A-317491, was antagonised by the coadministration of an anti-enkephalin antibody supporting the participation of endogenous enkephalins. Consistent with this result, the antihyperalgesic effect induced by A-317491 was dramatically enhanced by the administration of an enkephalin-degrading inhibitor, Debio 0827, as demonstrated by isobolographic analysis. This synergism opens the theoretical possibility that the combination of both types of drugs could be useful to counteract some nociceptive symptoms derived from tumor development.