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The Arve river valley airshed in the French Alps experiences particularly severe air pollution during wintertime stable atmospheric conditions associated with persistent cold-air pools. PM10 data recorded in the region indicate that the urbanised area of the central basin-shape section of the valley is generally the most polluted, with a harmful impact on the health of inhabitants. In the present work, we examine the air pollution transport potential of the Arve river valley airshed using results from high-resolution numerical simulations of a cold-air pool documented as part of the Passy-2015 field campaign. Passive tracers were used to model PM10 with emissions provided by a detailed inventory developed by the local air-quality agency. The observed differential in PM10 levels between valley sections was well captured by the numerical model and could not be explained solely by the differential in emissions. The stagnation, recirculation and ventilation potential of the airshed was evaluated spatially and temporally using integral quantities. The analysis indicated that the central basin-shape section of the valley is poorly ventilated and hence air pollution there would originate mostly from local emission sources. This stagnation zone appears to be almost decoupled from the rest of the airshed. The airshed was decomposed in separate valley sections so as to quantify the fate of the pollutants emitted within each section. Air pollution apportioned according to the contribution of emissions from the different valley sections shows that indeed the central basin-shape section is dominated by local sources. The situation was found more complex in the valley sections further downstream, where the contribution from the sum of the non-local sources can be as large as that from local sources. This study allows to identify the origin of the strong pollution in the Arve river valley, through the link between the local topography, emission sources and pollutant transport.

Temperature effects are of primary importance for designing concrete structures. Some of the early age temperature effects on the concrete behaviour can be accurately taken into consideration by well-known maturity functions. However, the effect of temperature on the autogenous deformations development is more complex, and results in contradictory evidence. This paper studies the influence of various isothermal curing temperatures from 10 °C to 30 °C on the autogenous deformations of concrete. Binary and ternary binders containing up to 30% of limestone filler and 70% of blast-furnace slag are studied. The amplitude of both the self-desiccation deformation and the early age swelling deformation are observed to decrease with increasing temperature, whatever the binder nature. Mechanisms for this observation are suggested, and a corresponding model is developed. The effect of the binder nature, age and temperature on the autogenous deformations is assessed with this model. Based on this new model, it is shown that the effect of temperature on the autogenous deformation development can be either beneficial or detrimental, depending on the nature of the binder., SCOPUS: ar.j, info:eu-repo/semantics/published

ispartof: Cement & Concrete Composites vol:85 pages:122-132 status: published

International audience; Background: Finite element modeling of the human head offers an alternative to experimental methods in understanding the biomechanical response of the head in trauma brain injuries. Falx, tentorium, and their notches are important structures surrounding the brain, and data about their anatomical variations are sparse. Objective: To describe and quantify anatomical variations of falx cerebri, tentorium cerebelli, and their notches. Methods: 3D reconstruction of falx and tentorium was performed by points identification on 40 brain CT-scans in a tailored Matlab program. A scatter plot was obtained for each subject, and 8 anatomical landmarks were selected. A reference frame was defined to determine the coordinates of landmarks. Segments and areas were computed. A reproducibility study was done. Results: The height of falx was 34.9 ± 3.9 mm and its surface area 56.5 ± 7.7 cm2. The width of tentorium was 99.64 ± 4.79 mm and its surface area 57.6 ± 5.8 cm2. The mean length, height, and surface area of falx notch were respectively 96.9 ± 8 mm, 41.8 ± 5.9 mm, and 28.8 ± 5.8 cm2 (range 15.8-40.5 cm2). The anterior and maximal widths of tentorial notch were 25.5 ± 3.5 mm and 30.9 ± 2.5 mm; its length 54.9 ± 5.2 mm and its surface area 13.26 ± 1.6 cm2. The length of falx notch correlated with the length of tentorial notch (r = 0.62, P < 0.05). Conclusion: We observe large anatomical variations of falx, tentorium, and notches, crucial to better understand the biomechanics of brain injury, in personalized finite element models.

Restrained deformations in concrete structures induce the development of stresses and sometimes cracking. Therefore the whole service life of concrete structures is influenced by the appropriate consideration of the early age behaviour of concrete under restrained conditions. For that reason, several devices were developed in the past to characterize the risk of cracking of cement-based materials. In the 1990s, a new experimental concrete testing concept has been designed: the Temperature Stress Testing Machine (TSTM). The device is able to monitor several parameters such as the age of cracking, the stiffness development and the stress relaxation. Between 1990 and 2010, less than fifteen laboratories worldwide have developed or acquired this kind of experimental equipment. All devices bear some similarities but major differences remain in the test set up designs, in the testing processes and also in the scale of the material used. This chapter summarizes all existing technics used to assess the behavior of concrete under restrained deformation, a comparison of the different existing TSTM is presented. New advanced techniques aim at monitor the cracking risk of cement-based materials such as the active ring test or the elliptical ring test are also presented.

General anesthesia reversibly alters consciousness, without shutting down the brain globally. Depending on the anesthetic agent and dose, it may produce different consciousness states including a complete absence of subjective experience (unconsciousness), a conscious experience without perception of the environment (disconnected consciousness, like during dreaming), or episodes of oriented consciousness with awareness of the environment (connected consciousness). Each consciousness state may potentially be followed by explicit or implicit memories after the procedure. In this respect, anesthesia can be considered as a proxy to explore consciousness. During the recent years, progress in the exploration of brain function has allowed a better understanding of the neural correlates of consciousness, and of their alterations during anesthesia. Several changes in functional and effective between-region brain connectivity, consciousness network topology, and spatio-temporal dynamics of between-region interactions have been evidenced during anesthesia. Despite a set of effects that are common to many anesthetic agents, it is still uneasy to draw a comprehensive picture of the precise cascades during general anesthesia. Several questions remain unsolved, including the exact identification of the neural substrate of consciousness and its components, the detection of specific consciousness states in unresponsive patients and their associated memory processes, the processing of sensory information during anesthesia, the pharmacodynamic interactions between anesthetic agents, the direction-dependent hysteresis phenomenon during the transitions between consciousness states, the mechanisms of cognitive alterations that follow an anesthetic procedure, the identification of an eventual unitary mechanism of anesthesia-induced alteration of consciousness, the relationship between network effects and the biochemical or sleep-wake cycle targets of anesthetic agents, as well as the vast between-studies variations in dose and administration mode, leading to difficulties in between-studies comparisons. In this narrative review, we draw the picture of the current state of knowledge in anesthesia-induced unconsciousness, from insights gathered on propofol, halogenated vapors, ketamine, dexmedetomidine, benzodiazepines and xenon. We also describe how anesthesia can help understanding consciousness, we develop the above-mentioned unresolved questions, and propose tracks for future research.

Intracerebroventricular (ICV) administration of opioids for control of intractable cancer pain has been used since 1982. We present here our experience of intracerebroventricular administration of pain treatments including ziconotide associated with morphine and ropivacaine for patients resistant to a conventional approach, with nociceptive, neuropathic, or mixed pain. These clinical cases were conducted with patients suffering from refractory pain, more than 6/10 on a numerical pain rating scale (NPRS) while on high-dose medical treatment and/ or intolerance with significant side effects from oral medication. The baseline study visit included a physical examination and an assessment of pain intensity on a NPRS. Under general anesthesia, a neuronavigation device was used to place the catheter on the floor of the third ventricle, supported by an endoscope. Then, drugs were injected in the cerebroventricular system, through a pump (external or subcutaneous). The primary objective was to measure pain evaluation with ICV treatment after a complete withdrawal of other medications. Four patients were enrolled: 3 with intractable cancer pain and one with central neuropathic pain. The median NPRS at baseline was 9.5 [8.5; 19]. The mean NPRS after one month was 3.5 [3; 4.5]. Ziconotide was initiated at 0.48 µg/dy and up to a median of 1.2 µg/dy [1.0; 1.56]. The median dose of morphine and ropivacaine used initially was respectively 0.36 mg/dy [0.24; 0.66] up to 0.6 mg/dy [0.45; 4.63] and 1.2 mg/dy [0; 2.4] up to 2.23 mg/dy [1.2; 3.35]. Minor side effects were initially observed but transiently. One psychiatric agitation required discontinuation of ziconotide infusion. For intractable pain, using ziconotide by intracerebroventricular infusion seems safe and efficient, specifically for chronic neoplastic pain of cervicocephalic, thoracic, or diffuse origin and also for pain arising from a central neuropathic mechanism. Key words: Intracerebroventricular infusion, ziconotide, intractable pain, nociceptive and neuropathic pain

The Stimulator of Interferon Genes (STING) protein is a key mediator of innate immunity that plays a central role in the immune response to invading pathogens (bacterial, viral) and transformed cells. A next generation human STING agonist, JNJ-‘6196 was developed that cures mice of their tumors in preclinical models when administered by the IV route. JNJ-‘6196 was rationally selected to have a weaker binding affinity and fast off rate but functionally is a strong cytokine inducer and an efficient activator of human dendritic cells. JNJ-‘6196 exhibits a unique cytokine induction profile in human PBMCs compared to other cyclic dinucleotides (CDNs) that are not curative by the IV route in mice with higher levels of pro-inflammatory cytokines that mediate antitumor activity and lower levels of those that promote suppressive M2 macrophages. In preclinical models of cancer in mice, JNJ-‘6196 eliminates bilateral tumors when administered IV, and demonstrates activity over a wide therapeutic range. Cured mice are immune to further re-challenge due to the expansion and persistence of tumor specific CD8+ T-cells following JNJ-‘6196 administration. Moreover, JNJ-‘6196 increased the effectiveness of checkpoint inhibitors, turning a PD-1 resistant model into a responsive model. Although it is a very potent inducer of antitumor cytokines in mouse and cyno, it is tolerated at similar dose levels as other CDNs that are not systemically active. The functional properties that confer systemic activity were investigated by comparing gene signatures of JNJ-‘6196 to another CDN that was not curative when administered by the IV route. Differences in the intensity of cytokine gene induction were likely responsible for systemic activity rather than genes that were selectively induced by this IV-active compound. The pharmacologic mode of action of JNJ-‘6196 was investigated and found to be Cmax driven based on efficacy and cytokine readouts. JNJ-‘6196 creates an immune inflamed microenvironment in tumors and could expand the population of patients that respond to immunotherapy. The ability to administer JNJ-'6196 systemically and the potential to synergize with other immunotherapeutics could create unique combination modalities and differentiate this compound from other STING agonists. Citation Format: Szeman Ruby Chan, Gilles Bignan, Emily Pierson, Sally Mahady, Hayley Ta, Wim Schepens, Jan Willem Thuring, Heng Keang Lim, Monicah Otieno, Thomas Wilde, Monica Singer, Nancy Bogdan, Shefali Patel, Leo Luistro, Liam Campion, Melissa Smith, Diana Wiley, Kathryn Packman, Michael Allegrezza, Caitlin Morgan, Jocelyn Sendecki, Glenn Van Aller, Daniel Krosky, Peter Connolly, James Edwards, Kim Staquet, Stuart L. Emanuel. JNJ-‘6196: A next generation STING agonist with potent preclinical activity by the IV route [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5567A.

This chapter reviews current knowledge of the effects of hypnotic anesthetic agents on brain resting-state networks (RSNs) that sustain consciousness. Although full exploration of the networks under anesthesia is not yet available, current evidence indicates that anesthetic agents with hypnotic properties dose-dependently modulate RSN functioning. Each anesthetic agent has specific effects that are not uniform within a given network and probably correlate with the specific clinical features observed when one agent or another is used. Observations made on RSNs during anesthesia are supplementary arguments to link the networks with specific aspects of consciousness and connectedness to the environment and to confirm their physiological functions. The precise link between observations made on RSNs during anesthesia and known biochemical targets of anesthetic agents, or their effects on systems that regulate the sleep–wake cycle, is not established yet. PET studies using radiolabeled probes that specifically target a neurotransmission system offer insights into the links. New technological advances and modes of functional data analysis, such as Granger causality and dynamic causal modeling, will help in obtaining a more in-depth exploration of the complex interactions between brain regions, their modulation by anesthesia, and their role in information processing by the brain. Effects of hypnosis on RSNs also have been studied. The hypnotic state is useful for performing surgical procedures and explorations without general anesthesia. The hypnotic state is associated with specific changes in the activity of RSNs that confirm hypnosis as a specific brain state, different from normal wakeful consciousness and anesthetic states.

Background Central neuropathic pain related to spinal cord injury is notoriously difficult to treat. So far most pharmacological and surgical options have shown but poor results. Recently ziconotide has been approved for use both neuropathic and non-neuropathic pain. In this cohort study, we assessed responder rate and long-term efficacy of intrathecal ziconotide in patients with pain related to spinal cord injury. Methods Patients presenting chronic neuropathic related to spinal cord lesions that was refractory to medical pain management were considered for inclusion. Those accepting were tested by lumbar puncture injection of ziconotide or continuous intrathecal infusion and if a significant decrease in pain scores (>40%) was noted they were implanted with a continuous infusion pump. They were then followed up for at least 1 year with constant assessment of the evolution of pain and side effects. Results Out of the 20 patients tested 14 had a decrease in pain scores of more than 40% but only 11 (55%) were implanted with permanent pumps due to side effects and patient choice. These were followed up on average for 3.59 years (±1.94) and in eight patients an above threshold decrease in pain scores was maintained. Overall in patients that responded to the test baseline VAS was 7.91 and 4.31 at last follow-up with an average dose of 7.2 μg of ziconotide per day. Six patients (30%) did not respond to any test and in three patients side effects precluded pump implantation. No significant long-term effects of the molecule were noted. Conclusion This study shows response to intrathecal ziconotide test in 40% of the patients of a very specific population in whom other therapeutic options are not available. This data justifies the development further studies such as a long-term randomized controlled trial. Significance Intrathecal Ziconotide is a posible alternative for the treatment of pain in patients with spinal cord injury and below level neuropathic pain.

Subgaleal hematoma in the newborn infant is rare, occurs early, and often bears serious consequences. We report on 2 subacute cases of bruising of the scalp that occurred following the use of a suction cup. Emergency treatment consisted of a transfusion of packed red blood cells and fresh frozen plasma. Children born by use of vacuum extractor or forceps require careful monitoring by the nursing staff throughout their stay in the maternity unit.

Summary Purpose of the study Anterior tarsectomy for pes cavus in adults is designed to relieve pain and correct the deformity. The present study reports radiological and clinical results with anterior tarsectomy in 39 cases of pes cavus. Material and methods The study concerned 39 cavus feet in 33 patients (22 males, 11 females; mean age: 31 years, range 16–49 years). Clinical outcome was assessed in terms of pain, function and motion, using the AOFAS classification. Radiological assessment (anteroposterior and lateral stress X-ray, views with Meary superficial wire-marking) measured the Djian angle, talometatarsal alignment, talar slope, calcaneal slope, calcaneal valgus, and osteoarthritis stage in adjacent joints. Results Mean follow-up was 9.8 years (range, 1–25). Mean AOFAS score at follow-up was 69.2/100 points (range, 14–100). Pain decreased considerably in 75% of cases, and 68% of patients recovered normal activity. The foot was aligned correctly in 67% of cases. At last follow-up, pes cavus remained undercorrected in 80% of feet, but mean Djian angle had improved from 100° to 111.3°. Calcaneal valgus improved from 30.8° to 24.8° and the podoscopic footprint was normal in 51% of feet. In 74% of feet, adjacent joints presented progressive osteoarthritic degeneration. Subjectively, 70% of patients were very satisfied or satisfied with minor reservations. Objective outcome was excellent or good in 66% of feet. Discussion and conclusion Outcome in terms of function, motion, complications and satisfaction was good, although pain relief results were poor. Anterior tarsectomy is able to correct initial pes cavus deformity and compensate anomalies of the hindfoot, but its correction capacity is limited, and its efficacy in case of clawfoot is poor. Anterior tarsectomy spares the adjacent Chopart complex and Lisfranc joints while inducing hypermobility, and leads to arthritis in 74% of cases. Better results are obtained in cases of reestablishment of the Meary-Tomeno line and of hindfoot valgus, as well as in cases of correction of equinus and clawfoot deformities. Worse results are observed in case of neurological evolutive disease or insufficient correction of the preceding deformities. Level of evidence: Level IV. Therapeutic Study.

The known cross-reactivity of monoclonal antibodies prepared against CD1 and HLA-DR antigens with skin components prompted us to study the reactivity pattern of human eccrine sweat glands with a panel of monoclonal antibodies directed against CD1 antigens (OKT6, BL6, D-47) and against HLA-class II antigens (anti-DR, BL2, LEU-10, IV-D12, MAJA-7). The labelling pattern of eccrine glands with the panel of monoclonal antibodies used in this study permits to establish three different antigenic compartments on eccrine glands: 1) acrosyringium and distal part of dermal duct anti-DR+, BL2+, LEU-10+, IV-D12+; 2) proximal part of dermal duct MAJA-7+; 3) secretory part D-47+. The immunological markers used in this work provide a useful tool for investigation of eccrine gland differentiation and human eccrine glandular pathology.

Resume Dans le traitement chirurgical du pied plat valgus reductible de l’adulte, l’arthrodese mediotarsienne a pour objectifs l’indolence et la correction de la deformation. Le but du travail etait l’evaluation radio-clinique des resultats de cette intervention. L’etude retrospective portait sur 22 pieds plats valgus reductibles chez 19 patients âges en moyenne de 43 ans (15-75). La revision clinique et radiographique etait realisee au recul moyen de 7 ans et 4 mois (6 mois-20 ans et 3 mois). Nous avons deplore deux pseudarthrodeses d’evolution favorable apres greffe spongieuse. A la revision, le score de Kitaoka etait de 73,5 points/100 (de 53 a 94). La douleur et la fonction etaient significativement ameliorees mais l’amplitude globale du pied en flexion/extension restait inchangee. Le pied etait axe dans 68 % des cas. La pente talienne et la divergence talo-calcaneenne etaient normalisees mais il persistait un defaut de correction de l’angle de Djian et une cassure de la ligne de Meary dans, respectivement, 68 et 41 % des cas. Le valgus calcaneen etait reduit de 6,6° (16,6° a 10°). Dans 50 % des cas, les articulations voisines presentaient une progression de la degenerescence arthrosique. Subjectivement, 73 % des patients etaient tres satisfaits et satisfaits. Objectivement, 68 % des resultats etaient excellents ou bons. Le recours a l’arthrodese mediotarsienne a permis une amelioration globale de la douleur ainsi qu’une recuperation fonctionnelle satisfaisante sans morbidite superieure a une simple arthrodese talo-naviculaire. Nos resultats restent, en ce sens, comparables a ceux de la litterature. Neanmoins, la restauration de la voute plantaire reste cliniquement et radiographiquement limitee. D’autre part, devant la frequence des remaniements arthrosiques, certes moderes et souvent asymptomatiques, des articulations voisines et face a la difficulte des patients jeunes a reprendre une activite sportive, les solutions chirurgicales conservatrices devraient certainement etre preferentiellement choisies.

Resume Le principe de stabilite primaire et secondaire de la prothese Esop repose sur un ancrage metaphysaire exclusif, non cimente. Cet implant modulaire est compose d’une metaphyse revetue d’hydroxyapatite, sur laquelle est adaptee en peroperatoire une piece diaphysaire jouant un role de simple centreur. L’objectif de cette etude retrospective continue etait d’evaluer les stabilites primaire et secondaire de l’implant Esop en mesurant la migration axiale au cours du temps. Entre 1995 et 2001, 172 PTH utilisant l’implant Esop et l’implant acetabulaire Atlas III, ont ete implantees dans notre service. Apres exclusion des patients decedes et perdus de vue nous avons revu 155 PTH chez 66 femmes et 32 hommes, d’âge moyen 57 ans, dont 53 % exercaient une activite professionnelle au moment de l’intervention. La coxarthrose et l’osteonecrose aseptique representaient 87 % des etiologies. Nous avons mesure a l’aide du logiciel Imagika la migration axiale et l’offset global des PTH a 4 periodes distinctes. Nous avons egalement analyse la survie et les resultats cliniques et radiographiques de cette serie grâce aux scores PMA. Le taux de survie des PTH de notre serie etait de 97,81 % (IC a 95 % 7,77 ; 8,03.), toutes causes confondues. Le score PMA retrouvait 97 % de resultats excellents, tres bons ou bons. Une migration de plus de 5 mm a ete mise en evidence dans 10 cas (6,4 %). Sur ces 10 cas, 7 avaient migre dans le premier mois, puis n’avaient pas evolue lors de la revision. L’offset femoral global etait diminue de plus de 10 mm dans 38 % des cas, temoignant de la medialisation du centre de rotation de la hanche. La stabilite primaire et secondaire de l’implant Esop est satisfaisante. La migration observee sur 10 implants correspondait dans 7 cas a un recalage lors de la remise en charge. Le centre de rotation de la hanche subissait globalement une medialisation : dans ces cas il serait interessant de disposer d’implants lateralises.

The generation of anti-variable region monoclonal antibodies (mAbs) against therapeutic antibodies is essential in the pharmacokinetic/pharmacodynamic (PK/PD) assessments of the drugs in clinical study samples. Sandwich EIA and other methods are typically employed to achieve sensitivity and selectivity for the PK/PD analyses. These assays usually require generation of mAb reagents that bind specifically to the therapeutic mAb candidate in non-competing pair combinations. Thus, large panels of anti-variable region mAbs must be generated in an expeditious manner to increase the probability of success. Previously, we described a novel immunization method using type 1 interferons (IFNs) coupled with an agonistic anti-CD40 mAb to drive immune responses (Staquet et al., Human Antibodies 15 (2006), 61-69). This protocol allows for rapid and robust generation of large panels of anti-variable region mAbs. In order to quickly characterize and efficiently identify optimal anti-variable region antibody pairs early in the hybridoma process using crude supernatants, an inexpensive, high-throughput ELISA method was developed. The ability to rapidly identify appropriate mAb pairs will save resources by eliminating the time-consuming and laborious process of subcloning irrelevant hybridomas.

Advanced distal neoplasia found at sigmoidoscopy could be the marker for more proximal lesions.In the setting of a screening clinic, subjects underwent flexible sigmoidoscopy. If no significant lesion was found, sigmoidoscopy was planned after 5 years. If an advanced neoplasia was found, colonoscopy was performed just after the first sigmoidoscopy and at 1, 3 and 5 years. If a non-advanced neoplasia was found, sigmoidoscopy was performed at 1, 3 and 5 years and followed by colonoscopy if advanced lesion was found.At first screening 1704/1912 (88%) subjects had a negative sigmoidoscopy, 104 (5.4%) had an advanced neoplasia, 96 (6%) had a non-advanced neoplasia and eight (0.4%) had invasive colorectal cancer (CRC). At follow-up examinations at 1, 3 and 5 years, among 170 subjects with advanced and non-advanced neoplasia, one developed invasive CRC and 47 (31.6%), advanced neoplasia. At 5 years, among 718 first negative sigmoidoscopies, 572 (80%) were confirmed negative and 97 (14%) had advanced neoplasia. Colorectal cancer status at 5 years could be checked for interval cancers in 97% of subjects and no CRC was diagnosed in subjects who did not attend sigmoidoscopy at 5 years. Comparison of the incidence of invasive CRC to the data of registries of the Netherlands and Luxembourg suggested that the incidence of CRC was decreased by 36%-46%. Seven of the nine CRCs were Duke's A and the two others were Duke's B and C.Screening with sigmoidoscopy followed by colonoscopy in case of positive sigmoidoscopy leads to substantial decreases in the incidence of CRC. Most CRCs found are at an early, curable stage of their development.

Integrin alphabeta heterodimers mediate adhesion to the extracellular matrix and at cell-cell contacts and initiate intracellular signalling cascades in response to a variety of inductive factors. Apart from the expression of alphavbeta3 that we have previously reported, little is known about the expression of integrins in odontoblasts. Here, we investigated the expression of alphav-binding beta integrin subunits in healthy human dental pulp in vivo and in odontoblasts differentiated in vitro. Reverse transcription/polymerase chain reaction analysis revealed the expression of alphav, beta1, beta5 and beta8 integrin mRNA, but not beta6, in whole pulp cells. Flow cytometry showed that the alphav and beta1 subunits were the most intensely expressed. Immunohistochemistry demonstrated that the beta1 subunit was localised in newly differentiated odontoblasts in the root and in mature odontoblasts in the crown, including their intradentinal cell processes. The alphav chain was predominantly expressed by mature odontoblasts and alphavbeta5 was only observed in mature odontoblasts. In vitro differentiated odontoblasts expressed genes for alphav, beta1 and beta5, but not for beta6 and beta8. A comparison of integrin profiles between cultured pulp cells and in vitro differentiated odontoblasts revealed that odontoblast maturation was characterised by a significant increase in the expression of alphav and beta1 subunits and alphavbeta5 integrin. The beta8 subunit was detected in nerve cells only. Histological analysis of teeth from alphav knockout mice showed no obvious structural modification in the odontoblast layer. Thus, human mature odontoblasts express alphavbeta3, alphavbeta5 and perhaps alphavbeta1 integrins, with the possible presence of alpha-beta1 pairs. The roles that these molecules play in the exchange of information throughout the odontoblast layer remain to be determined.

An ileal perforation occurred shortly after birth in 4 very premature newborns. Diagnosis was made on an abdominal distension with a pneumoperitoneum on X-ray. There were no biological, radiological nor histological signs of necrotizing enterocolitis. There were no digestive short- or long-term complications. According to the few authors who described this syndrome, there are some risk factors, but they were not clearly involved in our cases. Ileal perforation in the absence of signs of necrotizing enterocolitis is rarely reported but should be well known because of its good prognosis.

In the present study, we analyzed the phenotypic alterations induced by several allergens on immature dendritic cells (DC), with the aim to develop a potential in vitro alternative for predicting the sensitizing potential of chemicals. DC were generated from human monocytes cultured in the presence of GM-CSF, IL-4 and TGF-beta1 and treated for 2 or 4 days with different chemicals. Surface marker expression (HLA-DR, CD1a, CD40, CD54, CD83, CD86, CCR7 and E-cadherin) was analyzed by flow cytometry. Results showed that a 2-day treatment with the representative allergens DNCB and NiSO(4) induced significant changes of most antigens while other chemicals such as balm of Peru (strong allergen), kathon (moderate allergen), cinnamic aldehyde (mild allergen) or the irritant SLS had no significant effect. In contrast, the 4-day treatment with allergens substantially improved the results. Indeed, despite a large variability according to the donors, the number of modified antigens was significantly higher with all the tested chemicals, except kathon, as compared to that observed with the irritant SLS. The present study indicates that, in this model, the screening of mild or moderate allergens requires both the consideration of many antigens and a prolonged time of incubation with the chemicals.

Integrins are heterodimeric transmembrane receptors which promote cell adhesion, thus contributing to the maintenance of tissue organization in both normal and pathological conditions. To characterize the way odontoblasts may interact with other cells and the extracellular matrix in human teeth, we studied expression of alpha v beta 3 integrin, a putative receptor for osteoadherin. We showed that alpha v beta 3 integrin expression was restricted to odontoblasts, blood vessels, and small rounded cells in sound and carious pulp. Odontoblast staining intensity increased from the apical to the cusp region. Osteoadherin staining was strong in the whole odontoblast layer (with a slight decrease in the cusp region) and in predentin. Odontoblasts differentiating in vitro were stained with the anti-alpha v beta 3 integrin antibody, first at the level of intercellular contacts, then throughout the cell membrane. These results suggest that the alpha v beta 3 integrin could play a role in interodontoblast adhesion and odontoblast binding to the surrounding predentin/dentin/pulp matrix, possibly through osteoadherin.

Background: Migration and maturation of epidermal dendritic cells, the Langerhans cells (LC), are central events in the initiation of the cutaneous immune response. LC migration from skin to draining lymph nodes is regarded as an indispensable step for the early phase of antigen-specific sensitization. Among the several agents which influence the ability of LC to migrate, previous studies have revealed that matrix metalloproteinases (MMPs) and protein kinase C (PKC) contribute to promoting LC migration. In this work, we studied the effect of two recently developed PKC and MMPs inhibitors of vegetable origin on the migration of in vitro activated LC. Methods: The migratory capacity of epidermal and in vitro generated LC was assessed using a reconstituted basement membrane assay (Matrigel), mimicking the prerequisite passage through the dermal-epidermal basement membrane on the way to the lymph nodes. Results: Contact with chemical allergens, Bandrowski’s base or 2,4-dinitrobenzenesulfonic acid (DNBS), triggered migration. In the presence of PKC inhibitors, D-erythro-sphingosine and OX100, or an inhibitor of MMPs, LU105, allergen-induced migration of LC was strongly decreased. The association between OX100 and LU105 was more efficient in modulating the migration of activated LC compared to each molecule tested separately. Conclusions: These results showed that PKC and MMPs inhibitors act in synergy to inhibit the migration of activated epidermal dendritic cells in vitro. They underscore the role of PKC and MMPs inhibitors and suggest they may be of relevance for therapeutically regulating epidermal dendritic cell migration in inflammatory dermatoses.

This study evaluated the potential of the skin as a non invasive route for RSV vaccination using two G protein-derived molecules, G2Na and G5 in mice. G2Na contains T and B-cell epitopes whether G5 is a pure B-cell epitope. In contrast to G5, G2Na coadministered with CT three times at 1 month interval onto 1cm of square area shaved skin, elicited a consistent serum anti-G2Na and anti-CT IgG response. The anti-G2Na IgG response was dominated by IgG1 isotype, an indirect marker of a Th2 type of response. Dramatic reduction and decrease of RSV titers in lung tissues and in the nasal tract, respectively, following intranasal virus challenge revealed biological relevance of the transcutaneous immunization in the context of RSV vaccine. These results suggest that the transcutaneous route may offer a promising potential for novel RSV vaccine strategy, simple, painless and economical.

Summary Outer membrane protein (Omp)A is highly represented and conserved in the Enterobacteriaceae family. Using a recombinant OmpA from Klebsiella pneumoniae (kpOmpA), we have analysed the interaction between this bacterial cell wall protein and human Langerhans cells (LC), the antigen-presenting cells of the epidermis and mucosa. We showed that biotinylated kpOmpA binds to human LC freshly isolated from epidermis. kpOmpA up-regulated MHC class II, CD86 and CCR7 expression, enhanced migration in response to macrophage inflammatory protein-3β (MIP-3β) through a reconstituted basement membrane mimicking the prerequisite passage through the dermal-epidermal basement membrane on the way to lymph nodes. The allostimulatory function of kpOmpA-treated LC was more potent than that of untreated cells. Even though the proportion of LC which binds kpOmpA was shown to vary between individuals, our data indicate that kpOmpA binds to and activates LC, and suggest that recognition of OmpA by LC may be an initiating event in the antibacterial host response.

Human cord blood CD34+ progenitors cultured in the presence of granulocyte-macrophage colony-stimulating factor (GM-CSF), tumor necrosis factor-alpha (TNF-alpha) and transforming growth factor-beta (TGF-beta) generate a heterogeneous population of dendritic cells (DC), including Langerhans cells (LC). This combination of cytokines has been shown to be crucial for differentiation into LC. After day 5 of culture, TNF-alpha has been maintained in the medium in most studies despite the observation of spontaneous maturation of LC after day 12. Five-day samples of in vitro differentiated LC were cultured in parallel with or without TNF-alpha. The absence of TNF-alpha was shown to: (1) slow down proliferation without triggering apoptotic cell death, (2) enhance the percentage of LC, (3) delay or abrogate the expression of CD83, CD86, HLA-DR and CD208 molecules, and (4) maintain endocytosis by receptor and macropinocytosis. The withdrawal of TNF-alpha abrogated the spontaneous synthesis of matrix metalloproteinases. At day 12, TNF-alpha-deprived LC were less efficient in allogeneic T cell activation than LC cultivated with TNF-alpha. These data indicate that the suppression of TNF-alpha after day 5 maintains cells in an immature state and provides a population with 80% of LC at day 12.

Interleukin (IL)-9 is known to regulate many cell types involved in T-helper type 2 responses classically associated with asthma, including B- and T-lymphocytes, mast cells, eosinophils and epithelial cells. In contrast, target cells mediating the effects of IL-9 in the lower respiratory tract remain to be identified. Therefore, the authors evaluated the activity of IL-9 on human alveolar macrophages (AM) from healthy volunteers. AM preincubated with IL-9 before lipopolysaccharide (LPS) stimulation exhibited a decreased oxidative burst, as previously shown with IL-4. The inhibitory effect of IL-9 was abolished by anti-hIL-9R alpha monoclonal antibody, and presence of IL-9 receptors on AM was demonstrated by immunofluorescence. Both IL-4 and IL-9 failed to modulate tumour necrosis factor-alpha, IL-8 and IL-10 release by LPS-stimulated AM. However, several observations suggested that IL-9 and IL-4 act through different mechanisms: 1) interferon-gamma antagonised the IL4- but not the IL-9-mediated inhibition of AM oxidative burst; 2) expression of CD14 was downregulated by IL-4 but not by IL-9 and 3) production of tumour growth factor-beta by activated AM was potentiated by IL-9 and not by IL4, and was required for the IL-9-mediated inhibition of AM oxidative burst. These observations provide additional information concerning the activity of interleukin-9 in the lung, related to inflammatory or fibrosing lung processes.

Signals regulating the traffic of Langerhans cell precursors from blood to the epidermis are not yet fully understood. The observations that TGF-beta1 is of unique importance in Langerhans cells (LC) ontogeny and that macrophage inflammatory protein-3alpha (MIP-3alpha) is able to attract LC within the epidermis, prompted us to study the effect of MIP-3alpha and TGF-beta1 on the migration of LC precursors. The migratory capacity of immature dendritic cells (DC) was assessed using a reconstituted basement membrane assay (Matrigel), mimicking the prerequisite passage through the dermal-epidermal basement membrane on the way into the epidermis. DC differentiated from cord blood CD34 cells in the presence of GM-CSF plus TNF-alpha were subjected to migration using modified Boyden chambers. Day-6 DC progenitors migrated in a dose-dependent fashion in response to MIP-3alpha, and CD1alpha+ LC precursors responded preferentially to the chemokine. Immature DC did not respond strongly to TGF-beta1 alone in migration assays, but up to 68% of the cells migrated in response to MIP-3alpha plus TGF-beta1. Among them, at least 50% expressed CD1a and E-cadherin and can be considered LC precursors. The allostimulatory function of these cells was significantly more potent than that which migrated in response to MIP-3alpha alone. Our results show that a significant proportion of immature DC is able to migrate through a dermal-epidermal basement membrane equivalent. In the presence of TGF-beta1, the DC which respond to MIP-3alpha have the phenotype and the functional capacity of epidermal LC. Our findings underline the role of MIP-3alpha and TGF-beta1 in attraction and localization of immature LC within the epidermis under normal conditions.

The events occurring during the penetration of melanoma cells through the dermal-epidermal junction, which is the first crucial step in the process of metastasis, are poorly understood, partly because no suitable tissue models exist. In the in vitro model reported here, two melanoma clones (T1C3, which generates lung metastases in experimental animals, and IC8, which does not) derived from a single parental cell line were co-seeded with normal allogenic keratinocytes onto acellular human de-epidermized dermis with preserved intact basement membrane and cultured for up to 1 month at an air-liquid interface. Histological, immunohistochemical and ultrastructural studies showed that melanoma cells from the metastatic clone (T1C3), but not from the non-metastatic clone (IC8), penetrated the dermal-epidermal junction to invade the dermis after 3 weeks of culture. Local invasion was associated with the dissolution of the native epidermal basement membrane collagens type IV and VII. Confocal laser scanning microscopy analysis demonstrated that numerous T1C3 cells were able to colonize the interstitial dermis and to rapidly penetrate empty dermal cavities. Our model represents a significant technical advance over others currently available since: (i) the organized three-dimensional architecture of the native dermal-epidermal junction is preserved; (ii) the active invasion process coincides with the dissolution of native components of the epidermal basement membrane, i.e. collagen types IV and VII; and (iii) the ability of melanoma cells to cross the dermal-epidermal junction correlates with their metastatic potential. This model provides a valuable tool for the study of the time-course of the cellular and molecular events that occur during the earliest steps of invasion in cutaneous melanoma. It also offers new opportunities to study the possible role of the keratinocyte environment in melanoma invasion.

BACKGROUND: Megakaryocyte polyploidization results from the lack of cytoplasmic separation while the nucleus keeps dividing. METHODS: To investigate the role of actin in the megakaryocyte polyploidization, three human cell lines with megakaryocytic properties (DAMI, HEL and K562) were incubated in the presence of cytochalasin B, an inhibitor of actin polymerization. These data were then compared with normal megakaryocytes. RESULTS: Compared with control conditions, cells cultured in the presence of cytochalasin B revealed an augmentation of cell size and ploidy and an arrest of cell proliferation. The expression of platelet membrane glycoproteins Ib, IIb/IIIa, IIIa and thrombospondin and transferrin receptors was augmented after treatment with cytochalasin B. Physiologically, the role of actin in inducing polyploidization could be related to an imbalance between G- and F-actins. To test this hypothesis, we measured G-, F- and total actin in cytochalasin B-treated cells. Actin was found to be increased significantly in cytochalasin B-treated DAMI and HEL cell lines. In contrast, the G/F-actin ratio was not affected by cytochalasin B. To confirm these actin changes in physiological megakaryocytopoiesis, G- and F-actin contents were then estimated in normal megakaryocytes. The G- and F-actin contents of megakaryocytes from eight normal patients exponentially decreased from 2 to 128n, whereas the total actin content per cell kept increasing. The G/F ratio was unaffected. CONCLUSION: Polyploidization of human megakaryocytes results from either a diminution of actin synthesis or an increased actin turnover, which in turn possibly abrogates the formation of the actin cleavage furrow in telophasis.

Megakaryocyte polyploidization is an advantageous and regulated mechanism that leads to an increase in platelet production. In megakaryocytic cell lines, polyploidization can be obtained by using colchicine, an inhibitor of the tubulin spindle. The nucleolar organizer regions (AgNORs) are parts of nucleolar DNA transcribed into ribosomal RNA and are detected by the silver-staining technique. Their number is proportional to protein synthesis.To estimate protein synthesis in polyploid megakaryocytes, AgNORs are measured in three cell lines with megakaryocyte properties (DAMI, HEL and K562) after a 4-day culture in the presence or absence of colchicine. The mean number of AgNORs per cell was 16+/-4 (mean+/-SEM), 24+/-3 and 14+/-3 for DAMI, HEL and K-562 cell lines respectively. The addition of colchicine (10 ng mL[-1]) significantly increased the number of AgNORs per cell (DAMI 556%, HEL 338% and K-562 300% of controls, P0.05 using the t-test). Moreover, the number of nucleoles per cell after the addition of colchicine was augmented significantly (DAMI 246%; HEL 237% and K-562 148% of controls, P0.05 using the t-test). The total protein content estimated by Bradford's method increased significantly to 226%, 215% and 304% of controls in DAMI, HEL and K562 respectively (P005 using the t-test). After treatment with colchicine, the endomitotic index (EI) [mean of (log2 DNA content expressed in N)-1] measured by flow cytometry (and reflecting ploidy) increased to 234%, 255% and 301%, respectively, in DAMI, HEL and K-562 cell lines (P0.05 using the t-test). Concomitantly, the number of AgNORs per unit of DNA increased in the DAMI and HEL cell lines (P0.05 using the t-test) from 48+/-8 and 39+/-5, respectively, to 79+/-11 and 61+/-10. In contrast, the number of nucleoles and the total protein content per endomitotic index were not affected by colchicine (P0.05 using the t-test), but the number of nucleoles per endomitotic index of DAMI cells was affected.The increase in the number of the NORs induced by an agent known to stimulate polyploidization of megakaryocytic cell lines suggests that polyploidization occurs by a proportional increase in protein synthesis per DNA unit.

In addition to T cell receptor triggering, activation of T cells requires co-stimulatory signals that have been shown to be mainly initiated through CD28. We analyzed the expression and function of the two ligands for CD28, B7-1 (CD80) and B7-2 (CD86), on human Langerhans cells (LC), the antigen-presenting cells from epidermis. Human LC freshly isolated from epidermis (fLC) expressed significant level of B7-2, which was increased upon a short culture in vitro. In contrast, B7-1 was undetectable on fLC but appeared at the cell surface after a 3-day culture in vitro. Pre-incubation of 18-h cultured LC with anti-B7-2 monoclonal antibodies (mAb) was sufficient to abrogate the binding of CTLA4-Ig fusion protein, while a combination of both mAb against B7-1 and B7-2 was necessary to obtain a complete inhibition of CTLA4-Ig binding on 3-day cultured LC, showing the absence of a third CTLA4 ligand. The function of B7-1 and B7-2 on human LC has been analyzed by adding mAb at the beginning of mixed epidermal cell lymphocyte reactions. Anti-B7-2 mAb and CTLA4-Ig, but not anti-B7-1 mAb, strongly inhibited allogeneic, as well as recall antigen-induced T cell proliferation supported by fLC or 3-day cultured LC. Collectively, these results demonstrate that B7-2 is the major ligand for CD28/CTLA4 at the LC surface and that it plays a crucial role in human LC co-stimulatory function with little, if any, dependence on B7-1 expression.

GB36, a mouse monoclonal antibody (mAb) raised against an epithelial antigen of the human trophoblast, reacts with the epithelial basement membrane of chorionic villi; it does not react with the invasive extravillous cytotrophoblast. Expression and characterization of the antigen of GB36 (designated GBA36) were investigated in normal keratinocytes by immunoprecipitation, immunofluorescence and immunoelectron microscopic studies. Immunoprecipitation experiments demonstrated that the proteins identified on keratinocytes by mAb GB36 and a rat mAb anti-integrin alpha 6 (GoH3) were the same. Using immunofluorescence and immunoelectron microscopic methods, GBA36 was localized on the cell membrane facing the epithelial basal lamina of basal keratinocytes. GBA36 distribution in benign and malignant skin tumours was evaluated by immunostaining methods (immunofluorescence and immunoperoxidase). Analysis of tumours revealed that whereas benign epithelial tumours and intradermal naevi displayed high levels of GBA36, the expression of this antigen decreased progressively in spinocellular and basal cell carcinomas, and in cutaneous melanomas in relation to invasiveness. During cell transformation, GBA36 undergoes quantitative alterations, and expression is down-regulated. Although the functional relevance of these changes remains unknown, the correlation of decreased GBA36 expression with tumour progression may indicate a role for altered integrin expression in tissue invasion by human skin carcinoma and melanoma.

To study the migration capacity of Langerhans cells in vitro, a reconstituted basement membrane invasion assay (Matrigel assay) was performed. Partially enriched human epidermal Langerhans cell suspensions (6-20%) were incubated with or without haptens [2,4,6-/trinitrobenzenesulfonic acid (TNBS) and fluorescein isothiocyanate (FITC)] or an irritant sodium lauryl sulfate (SLS) in Hanks' balanced salt solution for 10 min at 37 degrees C, and were then used for the invasion assay. Fibroblast-conditioned medium prepared from human dermal fibroblasts was used as a source of chemoattractants. Non-treated cells showed a gradual increase in migration of HLA-DR-positive cells through the Matrigel-coated filters in a time-dependent manner. In vitro TNBS and FITC treatment resulted in a significant stimulation of this migration. By contrast, SLS treatment did not increase the number of migrating cells. The migration capacity of hapten-treated cells decreased both in the absence of human fibroblast-conditioned medium and in the presence of this medium in the upper compartment of the chamber, indicating that this medium actually functioned as chemoattractant. Our findings suggest that the contact between hapten and Langerhans cells could be one of the triggers to initiate and/or stimulate Langerhans cell migration from the epidermis to the dermis. In addition, some factors derived from the dermal fibroblasts may promote and regulate the directional migration of Langerhans cells.

Summary Background Computed tomography (CT) coupled with arthrography remains the reference standard for the preoperative evaluation of rotator cuff tears. The objectives of this study were to evaluate intra-observer and inter-observer reproducibilities of CT-arthrographic assessment of the subscapularis tendon and to assess the validity and reliability of this investigation. Hypothesis CT-arthrography is reliable and reproducible for the preoperative characterisation of subscapularis tendon lesions. Material and methods We retrospectively reviewed 67 shoulders with rotator cuff tears in one or more tendons managed by arthroscopy, both to confirm the diagnosis and to allow therapeutic interventions. Each of the 67 preoperative CT-arthrograms was evaluated by three readers, of whom the first two evaluated the images twice at an interval of 30 days. The following were recorded at each reading: partial- or full-thickness tear in one or more rotator cuff tendons, intra-tendinous delamination, tendon stump retraction, and fatty degeneration of the muscles. The position of the long head of biceps tendon was assessed. A statistical analysis was performed using Fleiss’ method to compute intra-observer and inter-observer variabilities in CT-arthrography assessment of the subscapularis tendon status. Validity of this assessment was measured by computing the concordance coefficients between CT-arthrography and arthroscopy. Results Specificity of CT-arthrography was satisfactory for assessing the subscapularis tendon. Sensitivity was low. Significant inter-observer and intra-observer variabilities were documented. The concordance coefficients between CT-arthrography and arthroscopy indicated that major differences were common with all three readers. When assessing the subscapularis tendon by CT-arthrography, all readers experienced difficulties in distinguishing intact tendons, delaminated tendons, and tendons with tears confined to the upper third. Discussion Although CT-arthrography remains the reference standard for the preoperative investigation of rotator cuff tears, significant variability occurs in assessing the continuity of the subscapularis tendon. Reliability of this assessment is not optimal, as shown by our evaluation of concordance with arthroscopy. Although our data should be interpreted in the light of the investigation and measurement biases present in our study, they suggest that CT-arthrography may fail to provide a valid and reproducible assessment of the subscapularis tendon. The development of magnetic resonance (MR) imaging and MR-arthrography will probably improve the preoperative evaluation of subscapularis tears in the near future. Level of evidence Level III; diagnostic value study.

Initial sensing of infection is mediated by germline-encoded pattern-recognition receptors (PRRs), the activation of which leads to the expression of inflammatory mediators responsible for the elimination of pathogens and infected cells. PRRs act as immune sensors that provide immediate cell responses to pathogen invasion or tissue injury. Here, we review the expression of PRRs in human dental pulp cells, namely, receptors from the Toll-like (TLR) and Nod-like NLR families, by which cells recognize bacteria. Particular attention is given to odontoblasts, which are the first cells encountered by pathogens and represent, in the tooth, the first line of defense for the host. Understanding cellular and molecular mechanisms associated with the recognition of bacterial pathogens by odontoblasts is critical for the development of therapeutic strategies that aim at preventing excessive pulp inflammation and related deleterious effects.

Human odontoblasts trigger immune response s to oral bacteria that invade dental tissues during the caries process. To date, their ability to regulate the expression of the nucleotide-binding domain leucine-rich repeat containing receptor NOD2 when challenged by Gram-positive bacteria is unknown. In this study, we investigated NOD2 expression in healthy and inflamed human dental pulps challenged by bacteria, and in cultured odontoblast-like cells stimulated with lipoteichoic acid (LTA), a Toll-like receptor (TLR) 2 agonist which is specific for Gram-positive bacteria. We found that NOD2 gene expression was significantly up-regulated in pulps with acute inflammation compared to healthy ones. In vitro, LTA augmented NOD2 gene expression and protein level in odontoblast-like cells. The increase was more pronounced in odontoblast-like cells compared to dental pulp fibroblasts. Blocking experiments in odontoblast-like cells with anti-TLR2 antibody strongly reduced the NOD2 gene expression increase, whereas stimulation with the synthetic TLR2 ligand Pam2CSK4 confirmed NOD2 gene up-regulation following TLR2 engagement. These data suggest that NOD2 up-regulation is part of the odontoblast immune response to Gram-positive bacteria and might be important in protecting human dental pulp from the deleterious effects of cariogenic pathogens.

Following autologous bone marrow transplantation (ABMT), both impaired T cell activation and defective production of the principal T cell growth factor, interleukin-2 (IL-2), has been observed. These processes are dependent on a rise of intracellular calcium ([Ca2+]i), a step which follows binding of T cell receptor (TCR) and transduction of signal via the generation of cytoplasmic second messengers. In order to better understand the nature of defective cellular immunity in ABMT, in the present study we investigated the rise of [Ca2+]i in T cells of recipients of ABMT. By concomitant labelling lymphocytes with anti-CD4 antibody and addition of fluo-3 as fluorescent calcium indicator, we have selected for the T cell subset which is the principal source of IL-2. Short-term (less than 1 year post-transplantation) recipients of ABMT show a statistically significant blunted rise in [Ca2+]i in response to concanavalin A as compared to normal controls not accounted for solely by a decreased percentage of CD4+ cells in these patients. The [Ca2+]i response of CD4+ cells from long-term (greater than 1 year post-transplant) recipients was lower than that of the normal group although not to a statistically significant level. These findings suggest that following ABMT is a defect in the early stages of T cell activation involving either T cell receptor binding or early signal transduction ultimately resulting in depressed transcription of IL-2 mRNA. These defects are analogous to findings in both allogeneic transplantation where factors of histoincompatibility and graft-versus-host disease (GVHD) come into play, as well as in the defective T cell activation of the normal ageing process.

A randomized, three-arm parallel study was performed to evaluate the acute analgesic potency and tolerance of a single oral dose of 40 mg piroxicam, 60 mg codeine, and a combination of 20 mg piroxicam and 30 mg codeine in 90 patients with chronic pain due to cancer. Thirty patients were randomized to each group. Pain intensity and pain relief were assessed at 1, 2, 3, 4, 5, and 6 hours after dosing using a four-point analog scale. At the end of the study period, both the patient and the observer gave a global impression of the efficacy of the therapy. Volunteered side effects and their severity were noted. The results indicated that the sum of the pain intensity differences was not statistically significant between the three groups (analysis of variance). For the overall evaluation of the efficacy, speed of action, and side effects, no statistically significant differences were seen between the three treatment groups. The results of this study suggest that piroxicam may be of value in the treatment of some types of cancer pain.

International audience; Recent studies have suggested that odontoblasts are involved in the dental pulp immune response to oral pathogens that invade human dentin during the caries process. How odontoblasts regulate the early inflammatory and immune pulp response to Gram-positive bacteria, which predominate in shallow and moderate dentin caries, is still poorly understood. In this study, we investigated the production of pro-and anti-inflammatory cytokines by odontoblast-like cells upon engagement of Toll-like receptor (TLR) 2, a pattern recognition molecule activated by Gram-positive bacteria components. We used a highly sensitive Milliplex (R) kit for detecting cytokines released by cells stimulated with lipoteichoic acid (LTA), a cell wall component of Gram-positive bacteria, or with the potent TLR2 synthetic agonist Pam2CSK4. We found that odontoblasts produce the pro-inflammatory cytokines interleukin (IL)-6 and CXCL8, as well as the immunosuppressive cytokine IL-10 in response to TLR2 agonists. GM-CSF, IFN gamma, IL-1 beta, IL-2, IL-4, IL-5, IL-7, IL-12(p70), IL-13 and TNF-alpha were not detected. These data indicate that TLR2 activation in human odontoblasts selectively induces production of mediators known to influence positively or negatively inflammatory and immune responses in pathogen-challenged tissues. We suggest that these molecules might be important in regulating the fine tuning of the pulp response to Gram-positive bacteria which enter dentin during the caries process. (C) 2010 Elsevier GmbH. All rights reserved.

We prepared and characterized golimumab (CNTO148), a human IgG1 tumor necrosis factor alpha (TNFα) antagonist monoclonal antibody chosen for clinical development based on its molecular properties. Golimumab was compared with infliximab, adalimumab and etanercept for affinity and in vitro TNFα neutralization. The affinity of golimumab for soluble human TNFα, as determined by surface plasmon resonance, was similar to that of etanercept (18 pM versus 11 pM), greater than that of infliximab (44 pM) and significantly greater than that of adalimumab (127 pM, p=0.018). The concentration of golimumab necessary to neutralize TNFα-induced E-selectin expression on human endothelial cells by 50% was significantly less than those for infliximab (3.2 fold; p=0.017) and adalimumab (3.3-fold; p=0.008) and comparable to that for etanercept. The conformational stability of golimumab was greater than that of infliximab (primary melting temperature [Tm] 74.8 °C vs. 69.5 °C) as assessed by differential scanning calorimetry. In addition, golimumab showed minimal aggregation over the intended shelf life when formulated as a high concentration liquid product (100 mg/mL) for subcutaneous administration. In vivo, golimumab at doses of 1 and 10 mg/kg significantly delayed disease progression in a mouse model of human TNFα-induced arthritis when compared with untreated mice, while infliximab was effective only at 10 mg/kg. Golimumab also significantly reduced histological scores for arthritis severity and cartilage damage, as well as serum levels of pro-inflammatory cytokines and chemokines associated with arthritis. Thus, we have demonstrated that golimumab is a highly stable human monoclonal antibody with high affinity and capacity to neutralize human TNFα in vitro and in vivo.

SummaryIntroductionAcromioclavicular dislocation (ACD) is frequent, at 8% of all shoulder traumas. Management in grade III lesions remains controversial. The present study assessed objective and subjective results at medium-term follow-up (mean, 60 months; range, 12–120 mo) in 27 patients managed by Ligastic® ligament reconstruction for acute dislocation.Patients and methodsThis is a multicenter, multi-surgeon retrospective study. Between 1998 and 2006, 59 patients were operated on for grade III or IV ACD, in one teaching hospital (Lille, France) and one general hospital (Tourcoing, France). Follow-up was performed by an independent (non-operator) observer. The 27 acute cases followed up underwent comparative bilateral radioclinical shoulder examination. Initial X-ray assessment found 14 grade-III (52%) and 13 grade-IV (48%) dislocations using Patte's classification.ResultsMean Constant score was significantly lower on the operated side (82.44 vs 90.04; p

[Introduction]: Map-1B belongs to the family of proteins that govern the dynamic state and organization of microtubules within cells. MAP-1B is a microtubule-associated protein highly expressed during the development of the nervous system. Its expression, regulated by the fragile X mental retardation protein (FMRP), is essential to stabilize microtubules during the elongation of dendrites and neurites. Other microtubules-associated molecules such as tau or MAP2 seem to act similarly. The aim of this work was to identify the MAP-1B expression in in vitro and in vivo human odontoblasts during development and carious processes. The expression of MAP2 and tau was also studied., [Materials and Methods]: In cultured cells, MAP-1B expression was analyzed by real-time polymerase chain reaction, flow cytometry, and Western blot. Its distribution was visualized by in situ hybridization and immunochemistry both in vitro and in vivo. The expression of FMRP, MAP2, and tau was identified by real-time polymerase chain reaction and immunochemistry., [Results]: MAP-1B is specifically expressed in odontoblasts from adult third molars as well as incisor germs from human embryos. In adult carious teeth, it is also expressed in newly differentiated dentin-forming cells. In vitro, MAP-1B expression is related to the differentiation state of odontoblasts. MAP-1B clearly underlines the cellular architecture of cell bodies and processes of differentiated cells. FMRP, MAP2, and tau are also detected in vivo., [Conclusion]: On the basis of these data, MAP-1B could be considered as a new protein involved in the terminal differentiation of odontoblasts.

Histiocytic cells infiltrating the lesions in eosinophilic granuloma of bone as well as in cutaneous histiocytosis X were studied using a murine monoclonal antibody (MA) produced with proliferating cells from an eosinophilic granuloma of bone. This MA reacts with Langerhans cells (LC) of normal human skin or mucous membranes and with proliferating cells of eosinophilic granuloma of bone and skin lesions of Letter-Siwe disease, as shown by immunohistochemistry and immunogold labelling. As other murine MA's obtained after immunization with human cortical thymocytes, this MA immunoprecipitates the 49-kDa CD1a antigen found on human LC and thymic-cell surfaces but not its breakdown product after treatment with trypsin, as demonstrated by analysis of immunoelectron labelling, cytofluorometry and gel electrophoresis. This first production of a CD1a MA from an eosinophilic granuloma supports the concept of Langerhans-cell histiocytosis.

The expression of VLA integrins on human epidermal Langerhans cells (LC) was investigated by indirect immunogold labeling on transmission electron microscopy, followed by a quantitative analysis. Labelings on suspensions enriched in freshly isolated LC were carried out with an antibody recognizing the beta 1 subunit, common to all members of the VLA family, and with antibodies specific for the six different alpha subunits, alpha 1 to alpha 6. Normal human epidermal LC were all beta 1 positive, of which 60% were highly positive. By labelings with different VLA-alpha-chain MoAb, there appeared two subpopulations of LC: one positive and one negative. On average 40% of LC bore small amounts of VLA-1 and VLA-3, 53% and 77% of LC expressed moderate amounts of VLA-2 and VLA-5, respectively, and VLA-4 and VLA-6 were expressed by 67% and 90% of LC, respectively. VLA proteins are mainly extracellular matrix protein receptors. VLA-6 (laminin receptor) and VLA-5 (fibronectin receptor) are expressed mainly by LC, and in this way could subserve LC to leave the epidermal compartment through the basement membrane, to migrate throughout the fibronectin network of the dermis before migrating via the afferent lymphatics to the regional lymph nodes, where they present antigen to T cells. VLA proteins such as VLA-4, VLA-3, or VLA-2, which are found involved in cell-cell contacts, could contribute to the promotion of T-cell activation by facilitating adherence between LC and T cells.

Herein we describe the use of an agonistic anti-murine CD40 MAb as a B cell proliferative agent to enhance the generation of monoclonal antibodies (MAbs) in Balb/c mice. While hybridoma technology has been validated repeatedly over the decades, little work has been described to improve upon the overall numbers of in vivo B cells and specific antibodies obtained from a fusion. To begin to address this situation, strategies to boost B lymphocyte yields for hybridoma production were employed. Anti-CD40 agonist antibodies have been reported to activate and amplify human resting B lymphocytes in vitro, resulting in increased cell numbers available for the generation of human hybridomas or B cell clones. An agonistic anti-murine CD40 MAb was administered to immunized mice 3 days prior to splenic harvest, and B lymphocyte yields were found to be approximately 2-fold higher in treated animals when compared to untreated animals. Moreover, the resulting hybridoma fusions using lymphocytes from treated animals yielded 5- to 10-fold more antigen reactive hybrids when compared to untreated animals. This novel addition to conventional approaches utilizes the proliferative effects of agonistic anti-CD40 MAbs to markedly enhance monoclonal antibody generation.

The generation of anti-variable region monoclonal antibodies (mAbs) against therapeutic antibodies is essential in the pharrnacokinetic/pharmacodynamic (PK/PD) assessments of the drugs in clinical study samples. Sandwich EIA and other methods are typically employed to achieve sensitivity and selectivity for the PK/PD analyses. These assays usually require generation of mAb reagents that bind specifically to the drug in non-competing pair combinations. Thus, large panels of anti-variable region mAbs must be generated in an expeditious manner to increase the probability of success. Herein we describe a novel immunization method that utilizes type 1 interferons (IFNs) as immunomodulators coupled with an agonistic anti-CD40 mAb as a B cell proliferative agent to drive immune responses. This novel protocol allows for rapid and robust mAb reagent generation without the use of conventional protein-denaturing adjuvants. The use of IFNs allowed for the generation of comparable and in some cases, increased numbers of anti-variable region mAbs in a dramatically shorter timeframe. This IFN based, immunostimulatory approach utilizing a non-denaturing adjuvant, likely presents conformational epitopes and may optimize the humoral response for the rapid generation of anti-variable region reagents to therapeutic mAb candidates.