Mutations in SLC1A4, encoding the brain serine transporter, are associated with developmental delay, microcephaly and hypomyelination

BACKGROUND L-serine plays an essential role in neuronal development and function. Although a non-essential amino acid, L-serine must be synthesised within the brain because of its poor permeability by the blood-brain barrier. Within the brain, its synthesis is confined to astrocytes, and its shuttle to neuronal cells is performed by a dedicated neutral amino acid transporter, ASCT1. METHODS AND RESULTS Using exome analysis we identified the recessive mutations, p.E256K, p.L315fs, and p.R457W, in SLC1A4, the gene encoding ASCT1, in patients with developmental delay, microcephaly and hypomyelination; seizure disorder was variably present. When expressed in a heterologous system, the mutations did not affect the protein level at the plasma membrane but abolished or markedly reduced L-serine transport for p.R457W and p.E256K mutations, respectively. Interestingly, p.E256K mutation displayed a lower L-serine and alanine affinity but the same substrate selectivity as wild-type ASCT1. CONCLUSIONS The clinical phenotype of ASCT1 deficiency is reminiscent of defects in L-serine biosynthesis. The data underscore that ASCT1 is essential in brain serine transport. The SLC1A4 p.E256K mutation has a carrier frequency of 0.7% in the Ashkenazi-Jewish population and should be added to the carrier screening panel in this community.

Active eosinophilic esophagitis is characterized by epithelial barrier defects and eosinophil extracellular trap formation.

BACKGROUND Eosinophilic esophagitis (EoE) exhibits esophageal dysfunction owing to an eosinophil-predominant inflammation. Activated eosinophils generate eosinophil extracellular traps (EETs) able to kill bacteria. There is evidence of an impaired barrier function in EoE that might allow pathogens to invade the esophagus. This study aimed to investigate the presence and distribution of EETs in esophageal tissues from EoE patients and their association with possible epithelial barrier defects. METHODS Anonymized tissue samples from 18 patients with active EoE were analyzed. The presence of DNA nets associated with eosinophil granule proteins forming EETs and the expression of filaggrin, the protease inhibitor lympho-epithelial Kazal-type-related inhibitor (LEKTI), antimicrobial peptides, and cytokines were evaluated by confocal microscopy following immune fluorescence staining techniques. RESULTS Eosinophil extracellular trap formation occurred frequently and was detected in all EoE samples correlating with the numbers of infiltrating eosinophils. While the expression of both filaggrin and LEKTI was reduced, epithelial antimicrobial peptides (human beta-defensin-2, human beta-defensin-3, cathelicidin LL-37, psoriasin) and cytokines (TSLP, IL-25, IL-32, IL-33) were elevated in EoE as compared to normal esophageal tissues. There was a significant correlation between EET formation and TSLP expression (P = 0.02) as well as psoriasin expression (P = 0.016). On the other hand, a significant negative correlation was found between EET formation and LEKTI expression (P = 0.016). CONCLUSION Active EoE exhibits the presence of EETs. Indications of epithelial barrier defects in association with epithelial cytokines are also present which may have contributed to the activation of eosinophils. The formation of EETs could serve as a firewall against the invasion of pathogens.

Morphological ripening of fluid inclusions and coupled zone-refining in quartz crystals revealed by cathodoluminescence imaging: Implications for CL-petrography, fluid inclusion analysis and trace-element geothermometry

Cathodoluminescence (CL) studies have previously shown that some secondary fluid inclusions in luminescent quartz are surrounded by dark, non-luminescent patches, resulting from fracture-sealing by late, trace-element-poor quartz. This finding has led to the tacit generalization that all dark CL patches indicate influx of low temperature, late-stage fluids. In this study we have examined natural and synthetic hydrothermal quartz crystals using CL imaging supplemented by in-situ elemental analysis. The results lead us to propose that all natural, liquid-water-bearing inclusions in quartz, whether trapped on former crystal growth surfaces (i.e., of primary origin) or in healed fractures (i.e., of pseudosecondary or secondary origin), are surrounded by three-dimensional, non-luminescent patches. Cross-cutting relations show that the patches form after entrapment of the fluid inclusions and therefore they are not diagnostic of the timing of fluid entrapment. Instead, the dark patches reveal the mechanism by which fluid inclusions spontaneously approach morphological equilibrium and purify their host quartz over geological time. Fluid inclusions that contain solvent water perpetually dissolve and reprecipitate their walls, gradually adopting low-energy euhedral and equant shapes. Defects in the host quartz constitute solubility gradients that drive physical migration of the inclusions over distances of tens of μm (commonly) up to several mm (rarely). Inclusions thus sequester from their walls any trace elements (e.g., Li, Al, Na, Ti) present in excess of equilibrium concentrations, thereby chemically purifying their host crystals in a process analogous to industrial zone refining. Non-luminescent patches of quartz are left in their wake. Fluid inclusions that contain no liquid water as solvent (e.g., inclusions of low-density H2O vapor or other non-aqueous volatiles) do not undergo this process and therefore do not migrate, do not modify their shapes with time, and are not associated with dark-CL zone-refined patches. This new understanding has implications for the interpretation of solids within fluid inclusions (e.g., Ti- and Al-minerals) and for the elemental analysis of hydrothermal and metamorphic quartz and its fluid inclusions by microbeam methods such as LA-ICPMS and SIMS. As Ti is a common trace element in quartz, its sequestration by fluid inclusions and its depletion in zone-refined patches impacts on applications of the Ti-in-quartz geothermometer.

A duplication in the canine beta-galactosidase gene GLB1 causes exon skipping and GM1-gangliosidosis in Alaskan huskies

GM(1)-gangliosidosis is a lysosomal storage disease that is inherited as an autosomal recessive disorder, predominantly caused by structural defects in the beta-galactosidase gene (GLB1). The molecular cause of GM(1)-gangliosidosis in Alaskan huskies was investigated and a novel 19-bp duplication in exon 15 of the GLB1 gene was identified. The duplication comprised positions +1688-+1706 of the GLB1 cDNA. It partially disrupted a potential exon splicing enhancer (ESE), leading to exon skipping in a fraction of the transcripts. Thus, the mutation caused the expression of two different mRNAs from the mutant allele. One transcript contained the complete exon 15 with the 19-bp duplication, while the other transcript lacked exon 15. In the transcript containing exon 15 with the 19-bp duplication a premature termination codon (PTC) appeared, but due to its localization in the last exon of canine GLB1, nonsense-mediated RNA decay (NMD) did not occur. As a consequence of these molecular events two different truncated GLB1 proteins are predicted to be expressed from the mutant GLB1 allele. In heterozygous carrier animals the wild-type allele produces sufficient amounts of the active enzyme to prevent clinical signs of disease. In affected homozygous dogs no functional GLB1 is synthesized and G(M1)-gangliosidosis occurs.

Pre-clinical in vivo models for the screening of bone biomaterials for oral/craniofacial indications: focus on small-animal models.

Preclinical in vivo experimental studies are performed for evaluating proof-of-principle concepts, safety and possible unwanted reactions of candidate bone biomaterials before proceeding to clinical testing. Specifically, models involving small animals have been developed for screening bone biomaterials for their potential to enhance bone formation. No single model can completely recreate the anatomic, physiologic, biomechanic and functional environment of the human mouth and jaws. Relevant aspects regarding physiology, anatomy, dimensions and handling are discussed in this paper to elucidate the advantages and disadvantages of small-animal models. Model selection should be based not on the 'expertise' or capacities of the team, but rather on a scientifically solid rationale, and the animal model selected should reflect the question for which an answer is sought. The rationale for using heterotopic or orthotopic testing sites, and intraosseous, periosseous or extraskeletal defect models, is discussed. The paper also discusses the relevance of critical size defect modeling, with focus on calvarial defects in rodents. In addition, the rabbit sinus model and the capsule model in the rat mandible are presented and discussed in detail. All animal experiments should be designed with care and include sample-size and study-power calculations, thus allowing generation of meaningful data. Moreover, animal experiments are subject to ethical approval by the relevant authority. All procedures and the postoperative handling and care, including postoperative analgesics, should follow best practice.

Loss of tapasin correlates with diminished CD8(+) T-cell immunity and prognosis in colorectal cancer.

BACKGROUND Tapasin is a crucial component of the major histocompatibility (MHC) class I antigen presentation pathway. Defects in this pathway can lead to tumor immune evasion. The aim of this study was to test whether tapasin expression correlates with CD8(+) cytotoxic T lymphocyte (CTL) infiltration of colorectal cancer (CRC) and overall survival. METHODS A next-generation tissue microarray (ngTMA) of 198 CRC patients with full clinicopathological information was included in this study. TMA slides were immunostained for tapasin, MHC I and CD8. Marker expression was analyzed with immune-cell infiltration, patient survival and TNM-staging. RESULTS A reduction of tapasin expression strongly correlated with venous invasion (AUC 0.682, OR 2.7, p = 0.002; 95% CI 1.7-5.0), lymphatic invasion (AUC 0.620, OR 2.0, p = 0.005; 95 % CI 1.3-3.3), distant metastasis (AUC 0.727, OR 2.9, p = 0.004; 95% CI 1.4-5.9) and an infiltrative tumor border configuration (AUC 0.621, OR 2.2, p = 0.017; 95% CI 1.2-4.4). Further, tapasin expression was associated with CD8(+) CTL infiltration (AUC 0.729, OR 5.4, p < 0.001; 95% CI 2.6-11), and favorable overall survival (p = 0.004, HR 0.6, 95% CI 0.42-0.85). CONCLUSIONS Consistent with published functional data showing that tapasin promotes antigen presentation, as well as tumor immune recognition and destruction by CD8(+) CTLs, a reduction in tapasin expression is associated with tumor progression in CRC.

Selective resistance to the PARP inhibitor olaparib in a mouse model for BRCA1-deficient metaplastic breast cancer

Metaplastic breast carcinoma (MBC) is a rare histological breast cancer subtype characterized by mesenchymal elements and poor clinical outcome. A large fraction of MBCs harbor defects in breast cancer 1 (BRCA1). As BRCA1 deficiency sensitizes tumors to DNA cross-linking agents and poly(ADP-ribose) polymerase (PARP) inhibitors, we sought to investigate the response of BRCA1-deficient MBCs to the PARP inhibitor olaparib. To this end, we established a genetically engineered mouse model (GEMM) for BRCA1-deficient MBC by introducing the MET proto-oncogene into a BRCA1-associated breast cancer model, using our novel female GEMM ES cell (ESC) pipeline. In contrast to carcinomas, BRCA1-deficient mouse carcinosarcomas resembling MBC show intrinsic resistance to olaparib caused by increased P-glycoprotein (Pgp) drug efflux transporter expression. Indeed, resistance could be circumvented by using another PARP inhibitor, AZD2461, which is a poor Pgp substrate. These preclinical findings suggest that patients with BRCA1-associated MBC may show poor response to olaparib and illustrate the value of GEMM-ESC models of human cancer for evaluation of novel therapeutics.

Periodontal regeneration - furcation defects: a consensus report from the AAP Regeneration Workshop

BACKGROUND Treatment of furcation defects is a core component of periodontal therapy. The goal of this consensus report is to critically appraise the evidence and to subsequently present interpretive conclusions regarding the effectiveness of regenerative therapy for the treatment of furcation defects and recommendations for future research in this area. METHODS A systematic review was conducted before the consensus meeting. This review aims to evaluate and present the available evidence regarding the effectiveness of different regenerative approaches for the treatment of furcation defects in specific clinical scenarios compared with conventional surgical therapy. During the meeting, the outcomes of the systematic review, as well as other pertinent sources of evidence, were discussed by a committee of nine members. The consensus group members submitted additional material for consideration by the group in advance and at the time of the meeting. The group agreed on a comprehensive summary of the evidence and also formulated recommendations for the treatment of furcation defects via regenerative therapies and the conduction of future studies. RESULTS Histologic proof of periodontal regeneration after the application of a combined regenerative therapy for the treatment of maxillary facial, mesial, distal, and mandibular facial or lingual Class II furcation defects has been demonstrated in several studies. Evidence of histologic periodontal regeneration in mandibular Class III defects is limited to one case report. Favorable outcomes after regenerative therapy for maxillary Class III furcation defects are limited to clinical case reports. In Class I furcation defects, regenerative therapy may be beneficial in certain clinical scenarios, although generally Class I furcation defects may be treated predictably with non-regenerative therapies. There is a paucity of data regarding quantifiable patient-reported outcomes after surgical treatment of furcation defects. CONCLUSIONS Based on the available evidence, it was concluded that regenerative therapy is a viable option to achieve predictable outcomes for the treatment of furcation defects in certain clinical scenarios. Future research should test the efficacy of novel regenerative approaches that have the potential to enhance the effectiveness of therapy in clinical scenarios associated historically with less predictable outcomes. Additionally, future studies should place emphasis on histologic demonstration of periodontal regeneration in humans and also include validated patient-reported outcomes. CLINICAL RECOMMENDATIONS Based on the prevailing evidence, the following clinical recommendations could be offered. 1) Periodontal regeneration has been established as a viable therapeutic option for the treatment of various furcation defects, among which Class II defects represent a highly predictable scenario. Hence, regenerative periodontal therapy should be considered before resective therapy or extraction; 2) The application of a combined therapeutic approach (i.e., barrier, bone replacement graft with or without biologics) appears to offer an advantage over monotherapeutic algorithms; 3) To achieve predictable regenerative outcomes in the treatment of furcation defects, adverse systemic and local factors should be evaluated and controlled when possible; 4) Stringent postoperative care and subsequent supportive periodontal therapy are essential to achieve sustainable long-term regenerative outcomes.

Deficiency of MALT1 paracaspase activity results in unbalanced regulatory and effector T and B cell responses leading to multiorgan inflammation

The paracaspase MALT1 plays an important role in immune receptor-driven signaling pathways leading to NF-κB activation. MALT1 promotes signaling by acting as a scaffold, recruiting downstream signaling proteins, as well as by proteolytic cleavage of multiple substrates. However, the relative contributions of these two different activities to T and B cell function are not well understood. To investigate how MALT1 proteolytic activity contributes to overall immune cell regulation, we generated MALT1 protease-deficient mice (Malt1(PD/PD)) and compared their phenotype with that of MALT1 knockout animals (Malt1(-/-)). Malt1(PD/PD) mice displayed defects in multiple cell types including marginal zone B cells, B1 B cells, IL-10-producing B cells, regulatory T cells, and mature T and B cells. In general, immune defects were more pronounced in Malt1(-/-) animals. Both mouse lines showed abrogated B cell responses upon immunization with T-dependent and T-independent Ags. In vitro, inactivation of MALT1 protease activity caused reduced stimulation-induced T cell proliferation, impaired IL-2 and TNF-α production, as well as defective Th17 differentiation. Consequently, Malt1(PD/PD) mice were protected in a Th17-dependent experimental autoimmune encephalomyelitis model. Surprisingly, Malt1(PD/PD) animals developed a multiorgan inflammatory pathology, characterized by Th1 and Th2/0 responses and enhanced IgG1 and IgE levels, which was delayed by wild-type regulatory T cell reconstitution. We therefore propose that the pathology characterizing Malt1(PD/PD) animals arises from an immune imbalance featuring pathogenic Th1- and Th2/0-skewed effector responses and reduced immunosuppressive compartments. These data uncover a previously unappreciated key function of MALT1 protease activity in immune homeostasis and underline its relevance in human health and disease.

Rescue of neonatal cardiac dysfunction in mice by administration of cardiac progenitor cells in utero

Striated preferentially expressed gene (Speg) is a member of the myosin light chain kinase family. We previously showed that disruption of the Speg gene locus in mice leads to a dilated cardiomyopathy with immature-appearing cardiomyocytes. Here we show that cardiomyopathy of Speg(-/-) mice arises as a consequence of defects in cardiac progenitor cell (CPC) function, and that neonatal cardiac dysfunction can be rescued by in utero injections of wild-type CPCs into Speg(-/-) foetal hearts. CPCs harvested from Speg(-/-) mice display defects in clone formation, growth and differentiation into cardiomyocytes in vitro, which are associated with cardiac dysfunction in vivo. In utero administration of wild-type CPCs into the hearts of Speg(-/-) mice results in CPC engraftment, differentiation and myocardial maturation, which rescues Speg(-/-) mice from neonatal heart failure and increases the number of live births by fivefold. We propose that in utero administration of CPCs may have future implications for treatment of neonatal heart diseases.

The Role of Cells in Meniscal Guided Tissue Regeneration: A Proof of Concept Study in a Goat Model.

OBJECTIVE Successful repair of defects in the avascular zone of meniscus remains a challenge in orthopedics. This proof of concept study aimed to investigate a guided tissue regeneration approach for treatment of tears in meniscus avascular zone in a goat model. DESIGN Full-depth longitudinal tear was created in the avascular zone of the meniscus and sutured. In the two treatment groups, porcine collagen membrane was wrapped around the tear without (CM) or with injection of expanded autologous chondrocytes (CM+cells), whereas in the control group the tear remained only sutured. Gait recovery was evaluated during the entire follow-up period. On explantation at 3 and 6 months, macroscopic gross inspection assessed healing of tears, degradation of collagen membrane, potential signs of inflammation, and osteoarthritic changes. Microscopic histology scoring criteria were developed to evaluate healing of tears, the cellular response, and the inflammatory response. RESULTS Gait recovery suggested protective effect of collagen membrane and was supported by macroscopical evaluation where improved tear healing was noted in both treated groups. Histology scoring in CM compared to suture group revealed an increase in tear margins contact, newly formed connective tissue between margins, and cell formations surrounded with new matrix after 3 months yet not maintained after 6 months. In contrast, in the CM+cells group these features were observed after 3 and 6 months. CONCLUSIONS A transient, short-term guided tissue regeneration of avascular meniscal tears occurred upon application of collagen membrane, whereas addition of expanded autologous chondrocytes supported more sustainable longer term tear healing.

Partial lateralization of the nasopalatine nerve at the incisive foramen for ridge augmentation in the anterior maxilla prior to placement of dental implants: a retrospective case series evaluating self-reported data and neurosensory testing

The objective of this study was to assess implant therapy after a staged guided bone regeneration procedure in the anterior maxilla by lateralization of the nasopalatine nerve and vessel bundle. Neurosensory function following augmentative procedures and implant placement, assessed using a standardized questionnaire and clinical examination, were the primary outcome variables measured. This retrospective study included patients with a bone defect in the anterior maxilla in need of horizontal and/or vertical ridge augmentation prior to dental implant placement. The surgical sites were allowed to heal for at least 6 months before placement of dental implants. All patients received fixed implant-supported restorations and entered into a tightly scheduled maintenance program. In addition to the maintenance program, patients were recalled for a clinical examination and to fill out a questionnaire to assess any changes in the neurosensory function of the nasopalatine nerve at least 6 months after function. Twenty patients were included in the study from February 2001 to December 2010. They received a total of 51 implants after augmentation of the alveolar crest and lateralization of the nasopalatine nerve. The follow-up examination for questionnaire and neurosensory assessment was scheduled after a mean period of 4.18 years of function. None of the patients examined reported any pain, they did not have less or an altered sensation, and they did not experience a "foreign body" feeling in the area of surgery. Overall, 6 patients out of 20 (30%) showed palatal sensibility alterations of the soft tissues in the region of the maxillary canines and incisors resulting in a risk for a neurosensory change of 0.45 mucosal teeth regions per patient after ridge augmentation with lateralization of the nasopalatine nerve. Regeneration of bone defects in the anterior maxilla by horizontal and/or vertical ridge augmentation and lateralization of the nasopalatine nerve prior to dental implant placement is a predictable surgical technique. Whether or not there were clinically measurable impairments of neurosensory function, the patients did not report them or were not bothered by them.

The influence of different fever definitions on the rate of fever in neutropenia diagnosed in children with cancer

BACKGROUND The temperature limit defining fever (TLDF) is based on scarce evidence. This study aimed to determine the rate of fever in neutropenia (FN) episodes additionally diagnosed by lower versus standard TLDF. METHODS In a single center using a high TLDF (39.0°C tympanic temperature, LimitStandard), pediatric patients treated with chemotherapy for cancer were observed prospectively. Results of all temperature measurements and CBCs were recorded. The application of lower TLDFs (LimitLow; range, 37.5°C to 38.9°C) versus LimitStandard was simulated in silicon, resulting in three types of FN: simultaneous FN, diagnosed at both limits within 1 hour; earlier FN, diagnosed >1hour earlier at LimitLow; and additional FN, not diagnosed at LimitStandard. RESULTS In 39 patients, 8896 temperature measurements and 1873 CBCs were recorded during 289 months of chemotherapy. Virtually applying LimitStandard resulted in 34 FN diagnoses. The predefined relevantly (≥15%) increased FN rate was reached at LimitLow 38.4°C, with total 44 FN, 23 simultaneous, 11 earlier, and 10 additional (Poisson rate ratioAdditional/Standard, 0.29; 95% lower confidence bound, 0.16). Virtually applying 37.5°C as LimitLow led to earlier FN diagnosis (median, 4.5 hours; 95% CI, 1.0 to 20.8), and to 53 additional FN diagnosed. In 51 (96%) of them, spontaneous defervescence without specific therapy was observed in reality. CONCLUSION Lower TLDFs led to many additional FN diagnoses, implying overtreatment because spontaneous defervescence was observed in the vast majority. Lower TLDFs led as well to relevantly earlier diagnosis in a minority of FN episodes. The question if the high TLDF is not only efficacious but as well safe remains open.

Transition bei angeborenen Herzfehlern in der Schweiz

Die Chance, mit einem angeborenen Herzfehler erwachsen zu werden, liegt mittlerweile bei über 90 %. Trotz aller Erfolge der Medizin können Herzfehler auch nach erfolgreicher Korrektur nicht als geheilt betrachtet werden. Langzeitkomplikationen im Sinn von Herzrhythmusstörungen, Herzinsuffizienz, embolischen Ereignissen, Endokarditis, oder Reoperationen aufgrund residualer Befunde oder Klappendegenerationen sind häufig. Aus diesem Grund wird für eine Vielzahl angeborener Herzfehler eine lebenslange spezialisierte Betreuung empfohlen. Jugendliche mit Herzfehlern müssen darauf vorbereitet werden, im Erwachsenenalter Eigenverantwortung zu übernehmen. Dazu ist eine strukturierte Transition ein entscheidender Vorteil. Dadurch werden einerseits Jugendliche selbst auf ihre Erwachsenenrolle vorbereitet und andererseits kann ein adäquater Transfer der Betreuung vom Kinder- zum Erwachsenenmediziner gewährleistet werden. Jugendliche ohne Transfer und ohne spezialisierte Betreuung im Erwachsenenalter haben ein erhöhtes Risiko, kardiovaskuläre Komplikationen zu erleiden. Es wurde gezeigt, dass die Mortalität bei angeborenen Herzfehlern bei Erwachsenen mit spezialisierter Betreuung geringer ist als bei Patienten ohne entsprechende Betreuung. Am Beispiel des Zentrums für angeborene Herzfehler in Bern wird aufgezeigt, wie eine pflegegeleitete Transitionssprechstunde aufgebaut und erfolgreich umgesetzt werden kann.