γδ T Cells Differentially Regulate Bone Loss in Periodontitis Models.

γδ T cells are nonclassical T lymphocytes representing the major T-cell population at epithelial barriers. In the gingiva, γδ T cells are enriched in epithelial regions adjacent to the biofilm and are considered to regulate local immunity to maintain host-biofilm homeostatic interactions. This delicate balance is often disrupted resulting in the development of periodontitis. Previous studies in mice lacking γδ T cells from birth (Tcrd-/- mice) examined the impact of these cells on ligature-induced periodontitis. Data obtained from those studies proposed either a protective effect or no impact to γδ T cells in this setting. Here, we addressed the role of γδ T cells in periodontitis using the recently developed Tcrd-GDL mice, enabling temporal ablation of γδ T cells. Specifically, the impact of γδ T cells during periodontitis was examined in 2 modalities: the ligature model and the oral infection model in which the pathogen Porphyromonas gingivalis was administrated via successive oral gavages. Ablation of γδ T cells during ligature-induced periodontitis had no impact on innate immune cell recruitment to the ligated gingiva. In addition, the number of osteoclasts and subsequent alveolar bone loss were unaffected. However, γδ T cells play a pathologic role during P. gingivalis infection, and their absence prevented alveolar bone loss. Further analysis revealed that γδ T cells were responsible for the recruitment of neutrophils and monocytes to the gingiva following the exposure to P. gingivalis. γδ T-cell ablation also downregulated osteoclastogenesis and dysregulated long-term immune responses in the gingiva. Collectively, this study demonstrates that whereas γδ T cells are dispensable to periodontitis induced by the ligature model, they play a deleterious role in the oral infection model by facilitating pathogen-induced bone-destructive immune responses. On a broader aspect, this study highlights the complex immunopathologic mechanisms involved in periodontal bone loss.

γδT Cells Are Essential for Orthodontic Tooth Movement.

Sustained mechanical forces applied to tissue are known to shape local immunity. In the oral mucosa, mechanical stress, either naturally induced by masticatory forces or externally via mechanical loading during orthodontic tooth movement (OTM), is translated, in part, by T cells to alveolar bone resorption. Nevertheless, despite being considered critical for OTM, depletion of CD4+ and CD8+ T cells is reported to have no impact on tooth movement, thus questioning the function of αßT cells in OTM-associated bone resorption. To further address the role of T cells in OTM, we first characterized the leukocytes residing in the periodontal ligament (PDL), the tissue of interest during OTM, and compared it to the neighboring gingiva. Unlike the gingiva, monocytes and neutrophils represent the major leukocytes of the PDL. These myeloid cells were also the main leukocytes in the PDL of germ-free mice, although at lower levels than SPF mice. T lymphocytes were more enriched in the gingiva than the PDL, yet in both tissues, the relative fraction of the γδT cells was higher than the αß T cells. We thus sought to examine the role of γδT cells in OTM. γδT cells residing in the PDL were mainly Vγ6+ and produced interleukin (IL)-17A but not interferon-γ. Using Tcrd-GDL mice allowing conditional ablation of γδT cells in vivo, we demonstrate that OTM was greatly diminished in the absence of γδT cells. Further analysis revealed that ablation of γδT cells decreased early IL-17A expression, monocyte and neutrophil recruitment, and the expression of the osteoclastogenic molecule receptor activator of nuclear factor-κß ligand. This, eventually, resulted in reduced numbers of osteoclasts in the pressure site during OTM. Collectively, our data suggest that γδT cells are essential in OTM for translating orthodontic mechanical forces to bone resorption, required for relocating the tooth in the alveolar bone.

Niche Specific Microbiota-Dependent and Independent Bone Loss around Dental Implants and Teeth.

Oral mucosal homeostasis is achieved by complex immunologic mechanisms, orchestrating host immunity to adapt to the physiologic functions of the various specialized niches in the oral cavity. Dental implants introduce a novel mucosal niche to the immune system to deal with. Nevertheless, the immune mechanisms engaged toward implants and whether they have broader effects are not well defined. Using a murine model, we found an accumulation of neutrophils and RANKL-expressing T and B lymphocytes in the implant-surrounding mucosa, accompanied by local bone loss. Surprisingly, the presence of implants had an impact on remote periodontal sites, as elevated inflammation and accelerated bone loss were detected in intact distant teeth. This was due to microbial dysbiosis induced by the implants, since antibiotic treatment prevented bone loss around teeth. However, antibiotic treatment failed to prevent the loss of implant-supporting bone, highlighting the distinct mechanisms mediating bone loss at each site. Further analysis revealed that implants induced chronic lymphocyte activation and increased mRNA expression of IFN-α and accumulation of IFN-α-producing plasmacytoid dendritic cells, which we previously reported as bone-destructive immune responses. Collectively, this study demonstrates that implants have a strong and broad impact on oral mucosal homeostasis, inducing periodontal bone loss in a niche-specific manner that is both microbiota dependent and independent.

Development and Function of γδT Cells in the Oral Mucosa.

To successfully withstand a wide variety of microbial and mechanical challenges, the immune system of the oral mucosa is composed of tissue-resident and specially recruited leukocytes. These leukocytes facilitate the establishment and maintenance of local homeostasis but are also capable to cause oral pathologies when are unrestrained. γδT cells represent an important tissue-resident innate T-cell population in various mucosal and nonmucosal barrier tissues, in which they are ideally located to assist in immunosurveillance, tissue repair, and homeostasis. Whereas most works studying γδT cells were focused on tissues such as the skin and intestine, these cells in the oral mucosa were only recently thoroughly studied. The findings obtained by those studies appear to be both complementary and contradicting, likely reflecting differences in the experimental settings and the type of transgenic mouse modalities employed by each study. Nevertheless, oral γδT cells were shown to consist of developmentally distinct tissue-resident Vγ6 cells and circulating Vγ1 and Vγ4 subsets that are independently maintained in the oral mucosa. In the gingiva, a particularly challenging barrier tissue due to its proximity to the dental plaque, γδT cells are strategically positioned close to the plaque and represent the major source of IL-17. While this suggests that γδT cells might be involved in controlling the dental biofilm, conflicting data were reported in this regard. In vivo studies have shown that γδT cells either play a protective role during age-associated bone loss or, alternatively, have no impact in this process. Also, recent reports suggested opposing data concerning the impact of γδT cells in experimental periodontitis based on the ligature model. This review summarizes and discusses the most up-to-date literature on oral γδT cells, providing a balanced perspective regarding our current understanding on the development of oral γδT cells and their role under physiologic conditions and certain oral pathologies.

T and B cell clonal expansion in Ras-associated lymphoproliferative disease (RALD) as revealed by next-generation sequencing.

Ras-associated lymphoproliferative disease (RALD) is an autoimmune lymphoproliferative syndrome (ALPS)-like disease caused by mutations in Kirsten rat sarcoma viral oncogene homologue (KRAS) or neuroblastoma RAS viral (V-Ras) oncogene homologue (NRAS). The immunological phenotype and pathogenesis of RALD have yet to be studied extensively. Here we report a thorough immunological investigation of a RALD patient with a somatic KRAS mutation. Patient lymphocytes were analysed for phenotype, immunoglobulin levels and T cell proliferation capacity. T and B cell receptor excision circles (TREC and KREC, respectively), markers of naive T and B cell production, were measured serially for 3 years. T and B cell receptor repertoires were studied using both traditional assays as well as next-generation sequencing (NGS). TREC and KREC declined dramatically with time, as did T cell receptor diversity. NGS analysis demonstrated T and B clonal expansions and marked restriction of T and B cell receptor repertoires compared to healthy controls. Our results demonstrate, at least for our reported RALD patient, how peripheral T and B clonal expansions reciprocally limit lymphocyte production and restrict the lymphocyte receptor repertoire in this disease. Decreased naive lymphocyte production correlated with a clinical deterioration in our patient's immune status, suggesting that TREC and KREC may be used as an aid in monitoring disease progression. Both the methodologies used here and the conclusions regarding immune homeostasis may be applicable to the research of ALPS and other immune dysregulation syndromes.

Microcephaly, intractable seizures and developmental delay caused by biallelic variants in TBCD: further delineation of a new chaperone-mediated tubulinopathy.

Microtubule dynamics play a crucial role in neuronal development and function, and several neurodevelopmental disorders have been linked to mutations in genes encoding tubulins and functionally related proteins. Most recently, variants in the tubulin cofactor D (TBCD) gene, which encodes one of the five co-chaperones required for assembly and disassembly of α/ß-tubulin heterodimer, were reported to underlie a recessive neurodevelopmental/neurodegenerative disorder. We report on five patients from three unrelated families, who presented with microcephaly, intellectual disability, intractable seizures, optic nerve pallor/atrophy, and cortical atrophy with delayed myelination and thinned corpus callosum on brain imaging. Exome sequencing allowed the identification of biallelic variants in TBCD segregating with the disease in the three families. TBCD protein level was significantly reduced in cultured fibroblasts from one patient, supporting defective TBCD function as the event underlying the disorder. Such reduced expression was associated with accelerated microtubule re-polymerization. Morpholino-mediated TBCD knockdown in zebrafish recapitulated several key pathological features of the human disease, and TBCD overexpression in the same model confirmed previous studies documenting an obligate dependency on proper TBCD levels during development. Our findings confirm the link between inactivating TBCD variants and this newly described chaperone-associated tubulinopathy, and provide insights into the phenotype of this disorder.

Evaluation of Fetal Weight Estimation Formulas in Assessing Small-for-Gestational-Age Fetuses.

PURPOSE: The aim of this study was to compare the accuracy of multiple sonographic fetal weight estimation models in assessing small-for-gestational-age (SGA) fetuses. MATERIALS AND METHODS: The cohort included all singleton pregnancies recorded at a single medical center from January 2004 to September 2011, with a minimum of 24 weeks of gestation. SGA was defined as a fetal weight of less than the 10th percentile. We used birth weight (BW) distribution curves for our population, matched according to fetal gender and gestational age. Predicted birth weights were calculated using 26 sonographic fetal weight estimation models, including targeted formulas for SGA fetuses. RESULTS: 1218 cases of SGA fetuses that underwent sonographic fetal weight estimation within one week prior to delivery were found. Prediction of fetal weight was significantly less accurate in SGA fetuses than in the general population. The random error for SGA fetuses ranged from 7.2 % to 13.9 % in different models, while the systematic error ranged from -12.8 % to 26 %. Most non-targeted formulas showed a specificity of over 90 % but a sensitivity of only 20 - 35 % in the detection of SGA fetuses, while most targeted formulas had a low specificity but a high sensitivity. The model by Scott et al. was found to be the most accurate in assessing SGA fetuses in our population. CONCLUSION: Estimation of fetal weight in SGA fetuses is less accurate than in the general population. Some formulas which are designed for SGA are more accurate than others and their use might increase the sensitivity in identifying SGA fetuses, with only a small decline in specificity.

SLC1A4 mutations cause a novel disorder of intellectual disability, progressive microcephaly, spasticity and thin corpus callosum.

Two unrelated patients, presenting with significant global developmental delay, severe progressive microcephaly, seizures, spasticity and thin corpus callosum (CC) underwent trio whole-exome sequencing. No candidate variant was found in any known genes related to the phenotype. However, crossing the data of the patients illustrated that they both manifested pathogenic variants in the SLC1A4 gene which codes the ASCT1 transporter of serine and other neutral amino acids. The Ashkenazi patient is homozygous for a deleterious missense c.766G>A, p.(E256K) mutation whereas the Ashkenazi-Iraqi patient is compound heterozygous for this mutation and a nonsense c.945delTT, p.(Leu315Hisfs*42) mutation. Structural prediction demonstrates truncation of significant portion of the protein by the nonsense mutation and speculates functional disruption by the missense mutation. Both mutations are extremely rare in general population databases, however, the missense mutation was found in heterozygous mode in 1:100 Jewish Ashkenazi controls suggesting a higher carrier rate among Ashkenazi Jews. We conclude that SLC1A4 is the disease causing gene of a novel neurologic disorder manifesting with significant intellectual disability, severe postnatal microcephaly, spasticity and thin CC. The role of SLC1A4 in the serine transport from astrocytes to neurons suggests a possible pathomechanism for this disease and implies a potential therapeutic approach.

A search for the most accurate formula for sonographic weight estimation by fetal sex - a retrospective cohort study.

OBJECTIVE: The aim of this study was to assess the effect of fetal sex on the accuracy of multiple formulas for sonographic estimation fetal weight (SEFW). METHODS: The cohort included all singleton live births recorded at a single medical center from January 2004 to September 2011. The accuracy of SEFW was compared between male and female fetuses using 6575 SEFW performed within 3 days prior to delivery. Fetal weight was estimated using 27 models. RESULTS: The accuracy of different formulas in predicting birth weight of male and female fetuses was found to be significantly different in almost every accuracy index that was compared (P < 0.05). The model by Sabbagha et al. was found to be the most accurate in assessing female fetuses. The most accurate model for male fetuses was a sex-specific formula by Melamed et al. We also found that a combination of the most accurate formula for each sex to one combined sex-specific model increased SEFW accuracy significantly. CONCLUSION: The accuracy of SEFW is significantly related to fetal sex. The combination of the formulas by Melamed et al. and Sabbagha et al. for male and female fetuses accordingly allowed more accurate SEFW in our research population.

Do assisted conception twins have an increased risk for anencephaly?

BACKGROUND: The incidence rates of anterior neural tube defects, anencephaly and encephalocele appear increased among twins compared with singletons. The current study aimed to evaluate whether the etiology of this phenomenon is related to twinning, assisted reproductive technology (ART), or both. METHODS: The study cohort consisted of parturient women who were referred to our ultrasonography unit between January 1998 and December 2009 due to suspicion of severe fetal abnormality. The study cohort was divided into two subgroups based on mode of conception: spontaneous and ART (including IVF and ICSI). The subgroups were further subdivided into singleton and multiple pregnancies. We also compared pregnancies diagnosed with anencephaly in the study group to all live births in the Department of Obstetrics and Gynecology. RESULTS: Anencephaly was diagnosed in 43 fetuses out of 1154 (3.7%) pregnancies diagnosed with severe fetal anomaly. Anencephaly was diagnosed in 9 out of 78 twin pregnancies (11.5%); of these, 8 of 45 (17.8%) were ART conceived and 1 of 33 (3%) spontaneously conceived. A significant correlation was found between twinning and anencephaly, with an odds ratio (OR) of 3.4 [confidence interval (CI) = 1.3-8.9, P= 0.011], while no significant correlation was found between ART and anencephaly. A significant correlation was found between anencephaly and the combination of ART conception and twinning (OR of 6.6, CI = 2.8-15.3, P< 0.01). Analyzing the distribution of pregnancies diagnosed with anencephaly in the study group compared with the total number of live births in the department revealed a significant correlation between twinning and anencephaly, with an OR of 11.4 (CI = 4.9-26.5, P< 0.01), with no significant correlation between ART and anencephaly. Among all live births, a significant correlation was found between anencephaly and the combination of ART conception and twinning (OR of 24.6, CI = 11.4-53.2, P< 0.01). CONCLUSIONS: Our data suggest that twin pregnancies conceived by ART constitute a high-risk group for anencephaly, due to a possible synergistic effect of twinning and ART.

Feasibility study of a novel approach to sore prevention in patients with spinal cord lesions: the computerized dynamic control Matrix 200 system.

OBJECTIVE: To evaluate the feasibility of a computerized mattress system based on a novel concept in sore prevention: continuous monitoring and adjustment of the interface pressure in small segments of contact between the skin and the supporting surface. DESIGN: A preliminary observational study. SETTING: The Spinal Department, Loewenstein Rehabilitation Hospital, Raanana, Israel. SUBJECTS: Twelve patients with spinal cord lesions. INTERVENTIONS: Patients were examined for signs of impending sores after lying on the mattress for up to 4 successive hours. The pressure within each of the mattress's air cells was continuously measured and adjusted. RESULTS: No evidence of redness or excessive perspiration was found in any of the areas considered to be high risk for bed sores. Maximal interface pressure was 22-30 mm Hg in most of the examinations. Most of the patients felt comfortable on the mattress and the staff adapted easily to its operation. CONCLUSIONS: The system is apparently safe, and at least as efficient as other existing means for preventing sores. In addition, it may allow for increased intervals between bed positionings. We conclude that this approach of pressure control has the potential to improve bed sore prevention in a rehabilitation hospital setting.