Porphyromonas gingivalis activates Heat-Shock-Protein 27 to drive a LC3C-specific probacterial form of select autophagy that is redox sensitive for intracellular bacterial survival in human gingival mucosa.

Porphyromonas gingivalis , a major oral pathobiont, evades canonical host pathogen clearance in human primary gingival epithelial cells (GECs) by initiating a non-canonical variant of autophagy consisting of Microtubule-associated protein 1A/1B-light chain 3 (LC3)-rich autophagosomes, which then act as replicative niches. Simultaneously, P. gingivalis inhibits apoptosis and oxidative-stress, including extracellular-ATP (eATP)-mediated reactive-oxygen-species (ROS) production via phosphorylating Heat Shock Protein 27 (HSp27) with the bacterial nucleoside-diphosphate-kinase (Ndk). Here, we have mechanistically identified that P. gingivalis -mediated induction of HSp27 is crucial for the recruitment of the LC3 isoform, LC3C, to drive the formation of live P. gingivalis -containing Beclin1-ATG14-rich autophagosomes that are redox sensitive and non-degrading. HSp27 depletions of both infected GECs and gingiva-mimicking organotypic-culture systems resulted in the collapse of P. gingivalis -mediated autophagosomes, and abolished P. gingivalis -induced LC3C-specific autophagic-flux in a HSp27-dependent manner. Concurrently, HSp27 depletion accompanied by eATP treatment abrogated protracted Beclin 1-ATG14 partnering and decreased live intracellular P. gingivalis levels. These events were only partially restored via treatments with the antioxidant N-acetyl cysteine (NAC), which rescued the cellular redox environment independent of HSp27. Moreover, the temporal phosphorylation of HSp27 by the bacterial Ndk results in HSp27 tightly partnering with LC3C, hindering LC3C canonical cleavage, extending Beclin 1-ATG14 association, and halting canonical maturation. These findings pinpoint how HSp27 pleiotropically serves as a major platform-molecule, redox regulator, and stepwise modulator of LC3C during P. gingivalis -mediated non-canonical autophagy. Thus, our findings can determine specific molecular strategies for interfering with the host-adapted P. gingivalis ' successful mucosal colonization and oral dysbiosis.

The Utility of Genetic Testing in Infantile Epileptic Spasms Syndrome: A Step-Based Approach in the Next-Generation Sequencing Era.

BACKGROUND: To evaluate the utility of genetic testing for etiology-specific diagnosis (ESD) in infantile epileptic spasms syndrome (IESS) with a step-based diagnostic approach in the next-generation sequencing (NGS) era. METHODS: The study cohort consisted of 314 patients with IESS, followed by the Pediatric Neurology Division of Ege University Hospital between 2005 and 2021. The ESD was evaluated using a step-based approach: step I (clinical phenomenology), step II (neuroimaging), step III (metabolic screening), and step IV (genetic testing). The diagnostic utility of genetic testing was evaluated to compare the early-NGS period (2005 to 2013, n = 183) and the NGS era (2014 to 2021, n = 131). RESULTS: An ESD was established in 221 of 314 (70.4%) infants with IESS: structural, 40.8%; genetic, 17.2%; metabolic, 8.3%; immune-infectious, 4.1%. The diagnostic yield of genetic testing increased from 8.9% to 41.7% in the cohort during the four follow-up periods. The rate of unknown etiology decreased from 34.9% to 22.1% during the follow-up periods. The genetic ESD was established as 27.4% with genetic testing in the NGS era. The genetic testing in the NGS era increased dramatically in subgroups with unknown and structural etiologies. The diagnostic yields of the epilepsy panels increased from 7.6% to 19.2%. However, the diagnostic yield of whole exome sequencing remained at similar levels during the early-NGS period at 54.5% and in the NGS era at 59%. CONCLUSIONS: The more genetic ESD (27.4%) was defined for IESS in the NGS era with the implication of precision therapy (37.7%).

Opportunistic pathogen Porphyromonas gingivalis targets the LC3B-ceramide complex and mediates lethal mitophagy resistance in oral tumors.

Mechanisms by which Porphyromonas gingivalis (P. gingivalis) infection enhances oral tumor growth or resistance to cell death remain elusive. Here, we determined that P. gingivalis infection mediates therapeutic resistance via inhibiting lethal mitophagy in cancer cells and tumors. Mechanistically, P. gingivalis targets the LC3B-ceramide complex by associating with LC3B via bacterial major fimbriae (FimA) protein, preventing ceramide-dependent mitophagy in response to various therapeutic agents. Moreover, ceramide-mediated mitophagy is induced by Annexin A2 (ANXA2)-ceramide association involving the E142 residue of ANXA2. Inhibition of ANXA2-ceramide-LC3B complex formation by wild-type P. gingivalis prevented ceramide-dependent mitophagy. Moreover, a FimA-deletion mutant P. gingivalis variant had no inhibitory effects on ceramide-dependent mitophagy. Further, 16S rRNA sequencing of oral tumors indicated that P. gingivalis infection altered the microbiome of the tumor macroenvironment in response to ceramide analog treatment in mice. Thus, these data provide a mechanism describing the pro-survival roles of P. gingivalis in oral tumors.

The evaluation of L-arginine solution as a solvent for propolis extraction: The phenolic profile, antioxidant, antibacterial activity, and in vitro bioaccessibility.

Ethanol has been widely used for the extraction of propolis. Due to its certain disadvantages, there has been an ongoing search to find alternative non-ethanolic extraction solvents. This study aimed to compare the phenolics, antioxidant, and antibacterial activity of propolis extracts prepared with 70% ethanol (EWE), propylene glycol (PGE), and L-arginine solution (BE). All extracts were subjected to an in vitro simulated digestion procedure, and the phenolic profile of non-digested and digested samples was determined by using LC-MS/MS. Additionally, the change in total phenolic (TPC), total flavonoid content (TFC), and antioxidant capacities were determined at each digestion phase. TPC and TFC of non-digested propolis extracts had similar values, although BE showed higher antioxidant capacity (p < .05). The amount of TPC reached or transformed at the intestinal stage was higher for BE and PG compared to EWE. BE also provided the highest antioxidant capacity assay in digested samples. The most common phenolics were pinocembrin, pinobanskin, galangin, and CAPE in non-digested extracts. However, their concentration was drastically reduced by digestion, and their recovery (R%) ranged from 0% to 9.38% of the initial amount detected in the non-digested extracts. Chrysin was the most bioaccessible flavonoid in all extracts. Among phenolic acids, the highest R% was determined for trans-cinnamic acid (22.14%) from BE. All extracts showed in vitro inhibitory activity against Escherichia coli and Staphylococcus aureus. This study suggests that an L-arginine solution could be used as an alternative solvent to ethanol and propylene glycol for propolis extraction.

Sprint and Anaerobic Power with the Soccer-Specific ACTN3 Gene: A Distintive Example

SUMMARY: The aim of this study is twofold: (1) to identify differences in certain anaerobic parameters (10m sprint, 30m sprint, anaerobic power, and Illinois agility tests) between professional and amateur soccer players, and (2) to determine whether there is a difference in the ACTN3 gene polymorphism between professional and amateur soccer players. Ultimately, the goal is to reveal which parameters contribute to the differentiation in these two aspects. A total of 133 volunteer soccer players, including 71 professionals and 62 amateurs, participated in the research. DNA extraction from buccal epithelial cells was performed using a commercial kit to determine the genetic background of the athletes, and Real-Time PCR was conducted for genotyping. Statistical analysis of the findings obtained from the test results was performed using the SPSS 23 (SPSS Inc., Chicago, IL, USA) package program. The homogeneity of variance of the data was assessed using the Levene Test, and normal distribution analyses were conducted using the Shapiro-Wilk Test. Chi-square and Mann-Whitney U tests were employed for parameter analysis. The significance level was set at p0.05). However, there is a statistically significant difference in anaerobic parameters (10m sprint, 30m sprint, and anaerobic power) except for the Illinois test (p<0.05). In conclusion, our study found that gene polymorphism is not a differentiating factor between professional and amateur soccer players, but speed (10m and 30m) and anaerobic power parameters are differentiating factors.

Los objetivos de este estudio fueron: 1º identificar diferencias en ciertos parámetros anaeróbicos (sprint de 10 m, sprint de 30 m, potencia anaeróbica y pruebas de agilidad de Illinois) entre jugadores de fútbol profesionales y amateurs, y 2º determinar si existe una diferencia en el polimorfismo del gen ACTN3 entre jugadores de fútbol profesionales y aficionados. En definitiva, el objetivo fue revelar qué parámetros contribuyen a la diferenciación en estos dos aspectos. En la investigación participaron un total de 133 jugadores de fútbol voluntarios, incluidos 71 profesionales y 62 aficionados. La extracción de ADN de las células epiteliales orales se realizó utilizando un kit comercial para determinar los antecedentes genéticos de los atletas y se realizó una PCR en tiempo real para el genotipado. El análisis estadístico de los hallazgos obtenidos a partir de los resultados de las pruebas se realizó utilizando el programa de paquete SPSS 23 (SPSS Inc., Chicago, IL, EE. UU.). La homogeneidad de la varianza de los datos se evaluó mediante la prueba de Levene y los análisis de distribución normal se realizaron mediante la prueba de Shapiro-Wilk. Para el análisis de parámetros se emplearon las pruebas de Chi-cuadrado y U de Mann-Whitney. El nivel de significancia se fijó en p0,05). Sin embargo, existe una diferencia estadísticamente significativa en los parámetros anaeróbicos (sprint de 10 m, sprint de 30 m y potencia anaeróbica) excepto para la prueba de Illinois (p<0,05). En conclusión, nuestro estudio encontró que el polimorfismo genético no es un fac- tor diferenciador entre jugadores de fútbol profesionales y amateurs, pero sí los parámetros de velocidad (10 m y 30 m) y potencia anaeróbica.

The role of multiple vitellogenins in early development of fishes.

Functions of vitellogenins have been in the limelight of fish reproductive physiology research for decades. The Vtg system of acanthomorph teleosts consists of two complete forms of Vtgs (VtgAa and VtgAb) and an incomplete form, VtgC. Insufficient uptake and processing of Vtgs and their yolk proteins lead to inadequate oocyte hydration ensuing failure in acquisition of egg buoyancy and early developmental deficiencies. This review presents a summary of our studies on utilization of multiple Vtgs in species with different egg buoyancy characteristics, as examples. Studies of moronids revealed limited degradation of all three forms of lipovitellin heavy chain derived from their three respective forms of Vtg, by which they contribute to the free amino acid pool driving oocyte hydration during oocyte maturation. In later studies, CRISPR/Cas9 was employed to invalidate zebrafish type I, type II and type III Vtgs, which are orthologs of acanthamorph VtgAa, VtgAb and VtgC, respectively. Results revealed type I Vtg to have essential developmental and nutritional functions in both late embryos and larvae. Genomic disturbance of type II Vtg led to high mortalities during the first 24 h of embryonic development. Despite being a minor form of Vtg in zebrafish and most other species, type III Vtg was also found to contribute essentially to the developmental potential of zebrafish zygotes and early embryos. Apart from severe effects on progeny survival, these studies also disclosed previously unreported regulatory effects of Vtgs on fecundity and fertility, and on embryo hatching. We recently utilized parallel reactions monitoring based liquid chromatography tandem mass spectrometry to assess the processing and utilization of lipovitellins derived from different forms of Vtg in Atlantic halibut and European plaice. Results showed the Lv heavy chain of VtgAa (LvHAa) to be consumed during oocyte maturation and the Lv light chain of VtgAb (LvLAb) to be utilized specifically during late larval stages, while all remaining YPs (LvLAa, LvHAb, LvHC, and LvLC) were utilized during or after hatching up until first feeding in halibut. In plaice, all YPs except LvHAa, which similarly to halibut supports oocyte maturation, are utilized from late embryo to late larval development up until first feeding. The collective findings from these studies affirm substantial disparity in modes of utilization of different types of Vtgs among fish species with various egg buoyancy characteristics, and they reveal previously unknown regulatory functions of Vtgs in maintenance of reproductive assets such as maternal fecundity and fertility, and in embryonic hatching. Despite the progress that has been made over the past two decades by examining multiple Vtgs and their functions, a higher complexity of these systems with much greater diversity between species in modes of Vtg utilization is now evident. Further research is needed to reveal novel ways each species has evolved to utilize these complex multiple Vtg systems and to discover unifying principles for this evolution in fishes of diverse lineages, habitats and life history characteristics.

Ultrasound-assisted deep eutectic solvent extraction of bioactive compounds from persimmon calyx.

This study aimed to investigate the ultrasound-assisted extraction of bioactive compounds from persimmon (Diospyros kaki) calyx by deep eutectic solvents (DES) with different molar ratios. For this reason, the prepared DES extracts' total phenolic-flavonoid compounds and antioxidant activities (1,1-diphenyl-2-picrilhydrazyl radical scavenging activity [DPPH•], Cupric Reducing Antioxidant Capacity (CUPRAC), and ferric reducing antioxidant power [FRAP]) were investigated as a result of the experimental design and optimization study conducted for this purpose. A sonication time of 20 min was determined as the optimal condition. Under these conditions, a molar ratio of 1.9:1 (lactic acid:choline chloride) and a water ratio of 70% provided the highest phenolic/flavonoid compounds and antioxidative activity. Correlations among water ratio, molar ratio, and sonication time were determined using principal component analysis (PCA). In conditions where total flavonoid compound, FRAP, and DPPH• are high due to PCA, it can be concluded that the sonication time is at high level; on the contrary, the water and molar ratios are at low level. In conclusion, ultrasound-assisted extraction using DES proved effective in persimmon calyx. Therefore, it can be recommended to use these environmentally friendly green solvents as an alternative to organic solvents in preparing extracts in various fields. PRACTICAL APPLICATION: This study shows the effectiveness of the ultrasound-assisted green extraction method using persimmon calyx specified as waste. These findings are compelling in the food industry in terms of consumers being now aware of green technology and the discovery that calyx is a good source of bioactive compounds.

Genomic disturbance of vitellogenin 2 (vtg2) leads to vitellin membrane deficiencies and significant mortalities at early stages of embryonic development in zebrafish (Danio rerio).

The specific functions and essentiality of type II vitellogenin (Vtg2) in early zebrafish development were investigated in this study. A vtg2-mutant zebrafish line was produced and effects of genomic disturbance were observed in F2 females and F3 offspring. No change in vtg2 transcript has been detected, however, Vtg2 abundance in F2 female liver was 5×, and in 1 hpf F3 vtg2-mutant embryos was 3.8× less than Wt (p < 0.05). Fecundity was unaffected while fertilization rate was more than halved in F2 vtg2-mutant females (p < 0.05). Hatching rate was significantly higher in F3 vtg2-mutant embryos in comparison to Wt embryos. Survival rate declined drastically to 29% and 18% at 24 hpf and 20 dpf, respectively, in F3 vtg2-mutant embryos. The introduced mutation caused vitelline membrane deficiencies, significant mortalities at early embryonic stages, and morphological abnormalities in the surviving F3 vtg2-mutant larvae. Overrepresentation of histones, zona pellucida proteins, lectins, and protein degradation related proteins in F3 vtg2-mutant embryos provide evidence to impaired mechanisms involved in vitellin membrane formation. Overall findings imply a potential function of Vtg2 in acquisition of vitellin membrane integrity, among other reproductive functions, and therefore, its essentiality in early zebrafish embryo development.

Use of polyglycolic acid-hyaluronic acid/ß-tricalcium phosphate scaffold with or without mesenchymal stem cells found ineffective in treating osteochondral lesions in rabbit knees.

OBJECTIVE: In this experimental animal study, a novel bilayered scaffold used in the treatment of osteochondral defects in rabbit knees was evaluated. This novel scaffold's upper (cartilage) layer consists of polyglycolic acid and hyaluronic acid, and the lower (bone) layer consists of ß-tricalcium phosphate. The purpose of this study was to evaluate the efficacy of this novel scaffold, combined with or without mesenchymal stem cells (MSCs), in the treatment of osteochondral defects in rabbit knees. METHODS: Osteochondral defects were created in the left femoral trochlea of 30 rabbits. In group A, defects were treated with scaffold combined with MSCs; in group B, defects were treated with cell-free scaffolds; and group C was a control group with defects left untreated. In the 12th week, animals were sacrificed for macroscopic evaluation. RESULTS: The mean International Cartilage Repair Society (ICRS) macroscopic scores were 4.95 for group A, 6.16 for group B, and 8.25 for group C. The mean Oswestry Arthroscopic Scores (OAS) were 1.65 for group A, 3.39 for group B, and 6.05 for group C. The macroscopic scores were significantly higher in group C than group A for ICRS scores and group A and group B for OAS (P < .001, P < .000, P < .022). CONCLUSION: In essence, our findings indicate that the newly developed osteochondral scaffold, when tested in a rabbit model, is not as effective as expected in repairing full-thickness osteochondral defects, with or without the supplementation of MSCs. Further investigation is required to enhance the effectiveness of this novel combination.

The Distinguishing Factor in Soccer Players is Aerobic Performance, not the ACTN3 Gene / El Factor Distintivo en los Jugadores de Fútbol es el Rendimiento Aeróbico y no el gen ACTN3

SUMMARY: The purpose of this study was to reveal the differences between ACTN3 genotype (RR, RX, XX) and aerobic performance [Yo-Yo IRT1 (m), VO2 max (ml/kg/min)] in professional and regional amateur league soccer players and to reveal which of these parameters was a distinctive factor in these athletes.71 professional soccer players (age: 23.66 ± 4.11 years; body height: 1.79 ± 6.99 m; body weight: 76.02 ± 6.76 kg; body fat: 11.59±3.11 %) and 62 regional amateur soccer players (age: 23.63 ±3.77 years; body height: 1.81 ± 5.77 m; body weight: 76.36 ± 7.53 kg; body fat: 15.60±4.65 %) volunteered for the study. After DNA extraction from buccal epithelial cells via a commercial kit was performed for the genetic background of the athletes, Real-Time PCR was carried out for genotyping. Furthermore, Yo-Yo IRT1 test was performed to determine the aerobic performance of the soccer players. SPSS 23 (SPSS Inc., Chicago, IL, USA) package program was used for the statistical analysis of the data obtained in the tests. Shapiro-Wilk test for normality and Levene's test for homogeneity of variance were performed. Chi-Square, Independent Sample T Test and One Way ANOVA test were used in the analysis of the parameters. Statistical significance was set as p0.05); however, there was a statistical significance in favor of professional soccer players in terms of aerobic parameters (p<0.05). Consequently, it can be said that aerobic performance is the distinguishing factor, not the ACTN3 gene, in soccer players.

El objetivo de este estudio fue revelar las diferencias entre el genotipo ACTN3 (RR, RX, XX) y el rendimiento aeróbico [Yo-Yo IRT1 (m), VO2 max (ml/kg/min)] en jugadores de fútbol de ligas profesionales y amateurs regionales y determinar cuál de estos parámetros es un factor distintivo en estos deportistas. 71 futbolistas profesionales (edad: 23,66 ±4,11 años; altura corporal: 1,79 ± 6,99 m; peso corporal: 76,02 ± 6,76 kg; grasa corporal: 11,59±3,11 %) y 62 jugadores de fútbol amateur regionales (edad: 23,63 ± 3,77 años; altura corporal: 1,81 ± 5,77 m; peso corporal: 76,36 ± 7,53 kg; grasa corporal: 15,60 ± 4,65 %) se ofrecieron como voluntarios para el estudio. Después de realizar la extracción de ADN de las células epiteliales orales mediante un kit comercial para obtener los antecedentes genéticos de los atletas, se llevó a cabo una PCR en tiempo real para el genotipado. Además, se realizó la prueba Yo-Yo IRT1 para determinar el rendimiento aeróbico de los futbolistas. Para el análisis estadístico de los datos obtenidos en las pruebas se utilizó el programa SPSS 23 (SPSS Inc., Chicago, IL, EE. UU.). Se realizó la prueba de normalidad de Shapiro- Wilk y la prueba de homogeneidad de la varianza de Levene. En el análisis de los parámetros se utilizaron Chi-cuadrado, prueba T para muestra independiente y prueba ANOVA unidireccional. La significancia estadística se estableció en p0,05); sin embargo, hubo significación estadística a favor de los futbolistas profesionales en cuanto a los parámetros aeróbicos (p<0,05). En consecuencia, se puede decir que el rendimiento aeróbico es el factor distintivo, no el gen ACTN3, en los jugadores de fútbol.

Physical activity and exercise for the prevention and management of mild cognitive impairment and dementia: a collaborative international guideline.

BACKGROUND: Physical activity and exercise have been suggested as effective interventions for the prevention and management of mild cognitive impairment (MCI) and dementia, but there are no international guidelines. OBJECTIVES: To create a set of evidence- and expert consensus-based prevention and management recommendations regarding physical activity (any bodily movement produced by skeletal muscles that results in energy expenditure) and exercise (a subset of physical activity that is planned, structured, repetitive), applicable to a range of individuals from healthy older adults to those with MCI/dementia. METHODS: Guideline content was developed with input from several scientific and lay representatives' societies. A systematic search across multidisciplinary databases was carried out until October 2021. Recommendations for prevention and management were developed according to the GRADE and complemented by consensus statements from the expert panels. RECOMMENDATIONS: Physical activity may be considered for the primary prevention of dementia. In people with MCI there is continued uncertainty about the role of physical activity in slowing the conversion to dementia. Mind-body interventions have the greatest supporting evidence. In people with moderate dementia, exercise may be used for maintaining disability and cognition. All these recommendations were based on a very low/low certainty of evidence. CONCLUSIONS: Although the scientific evidence on the beneficial role of physical activity and exercise in preserving cognitive functions in subjects with normal cognition, MCI or dementia is inconclusive, this panel, composed of scientific societies and other stakeholders, recommends their implementation based on their beneficial effects on almost all facets of health.

The relationships between ACTN3 rs1815739 and PPARA-α rs4253778 gene polymorphisms and athletic performance characteristics in professional soccer players.

BACKGROUND: Current research on athletic performance focuses on genetic variants that contribute significantly to individuals' performance. ACTN3 rs1815739 and PPARA-α rs4253778 gene polymorphisms are variants frequently associated with athletic performance among different populations. However, there is limited research examining the pre-and post-test results of some variants of athletic performance in soccer players. Therefore, the presented research is to examine the relationships between the ACTN3 rs1815739 and PPARA-α rs4253778 gene polymorphisms and athletic performance improvement rates in adaptations to six weeks of training in elite soccer players using some athletic performance tests. METHODOLOGY: Twenty-two soccer players between the ages of 18 and 35 voluntarily participated in the study. All participants were actively engaged in a rigorous six-day-a-week training program during the pre-season preparation period. Preceding and following the training program, a battery of diverse athletic performance tests was administered to the participants. Moreover, Genomic DNA was extracted from oral epithelial cells using the Invitrogen DNA isolation kit (Invitrogen, USA), following the manufacturer's protocol. Genotyping was conducted using real-time PCR. To assess the pre- and post-test performance differences of soccer players, the Wilcoxon Signed Rank test was employed. RESULTS: Upon analyzing the results of the soccer players based on the ACTN3 genotype variable, it was observed that there were no statistically significant differences in the SJ (Squat Jump), 30m sprint, CMJ (Counter Movement Jump), and DJ (Drop Jump) performance tests (p > 0.05). However, a statistically significant difference was identified in the YOYO IRT 2 (Yo-Yo Intermittent Recovery Test Level 2) and 1RM (One Repetition Maximum) test outcomes (YOYO IRT 2: CC, CT, and TT, p = 0.028, 0.028, 0.008, 0.000, respectively; 1RM: CC, CT, and TT, p = 0.010, 0.34, 0.001, respectively). Regarding the PPARA-α genotype variable, the statistical analysis revealed no significant differences in the SJ, 30m sprint, CMJ, and DJ performance tests (p > 0.05). Nevertheless, a statistically significant difference was observed in the YOYO IRT 2 and 1RM test results (YOYO IRT 2: CC, CG p = 0.001, 0.020; 1RM: CC, p = 0.000) CONCLUSIONS: The current study demonstrated significant enhancements in only YOYO INT 2 and 1RM test outcomes across nearly all gene variants following the six-day-a-week training program. Other performance tests, such as the 30m sprint, SJ, CMJ, and DJ tests did not exhibit statistically significant differences. These findings contribute novel insights into the molecular processes involving PPARA-α rs4253778 and ACTN3 rs1815739 that underpin enhancements in endurance (YOYO INT 2) and maximal strength (1RM) aspects of athletic performance. However, to comprehensively elucidate the mechanisms responsible for the association between these polymorphisms and athletic performance, further investigations are warranted. It is thought that the use of field and genetic analyses together to support each other will be an important detail for athletes to reach high performance.

Statins Modulate Microenvironmental Cues Driving Macrophage Polarization in Simulated Periodontal Inflammation.

Periodontal disease (PD) is a chronic inflammatory disorder characterized by the destruction of connective tissue, tooth loss, and systemic infections. Clinically, treatment of PD includes control of the etiologic factors via several modalities: initial therapy including scaling and root planing (SRP), corrective phase of surgical treatment, both with and without adjunct antimicrobial/pharmacological agents, followed by a maintenance/supportive periodontal therapy phase. Each treatment phase aims to control oral biofilm by addressing risk factors and etiology. Monotherapy of systemic antibiotics is insufficient compared to their use as an adjunct to SRP. The critical issue of systemic antimicrobial usage includes adverse patient outcomes and increased bacterial resistance. Therefore, alternative adjuncts to periodontal therapy have been sought. Statins are widely prescribed for the treatment of hypercholesterolemia and cardiovascular disease. Statins have demonstrated anti-inflammatory properties and immunomodulatory effects, and a few retrospective studies showed that statin patients exhibit fewer signs of periodontal inflammation than subjects without the medication. Despite the available clinical studies on the local administration of statins for PD, no studies have reported the macrophage polarization response. We have developed a gingival fibroblast-macrophage co-culture model to track macrophage response when exposed to a battery of microenvironmental cues mimicking macrophage polarization/depolarization observed in vivo. Using our model, we demonstrate that simvastatin suppresses macrophage inflammatory response and upregulates tissue homeostasis and M2 macrophage markers. Our findings support the usage of statins to mitigate periodontal inflammation as a valid strategy.

The Presence of Periodontitis Exacerbates Non-Alcoholic Fatty Liver Disease via Sphingolipid Metabolism-Associated Insulin Resistance and Hepatic Inflammation in Mice with Metabolic Syndrome.

Clinical studies have shown that periodontitis is associated with non-alcoholic fatty liver disease (NAFLD). However, it remains unclear if periodontitis contributes to the progression of NAFLD. In this study, we generated a mouse model with high-fat diet (HFD)-induced metabolic syndrome (MetS) and NAFLD and oral P. gingivalis inoculation-induced periodontitis. Results showed that the presence of periodontitis increased insulin resistance and hepatic inflammation and exacerbated the progression of NAFLD. To determine the role of sphingolipid metabolism in the association between NAFLD and periodontitis, we also treated mice with imipramine, an inhibitor of acid sphingomyelinase (ASMase), and demonstrated that imipramine treatment significantly alleviated insulin resistance and hepatic inflammation, and improved NAFLD. Studies performed in vitro showed that lipopolysaccharide (LPS) and palmitic acid (PA), a major saturated fatty acid associated with MetS and NAFLD, synergistically increased the production of ceramide, a bioactive sphingolipid involved in NAFLD progression in macrophages but imipramine effectively reversed the ceramide production stimulated by LPS and PA. Taken together, this study showed for the first time that the presence of periodontitis contributed to the progression of NAFLD, likely due to alterations in sphingolipid metabolism that led to exacerbated insulin resistance and hepatic inflammation. This study also showed that targeting ASMase with imipramine improves NAFLD by reducing insulin resistance and hepatic inflammation.

Rottlerin and genistein inhibit neuroblastoma cell proliferation and invasion through EF2K suppression and related protein pathways.

Neuroblastoma is one of the most common solid tumors in children younger than 1 year of age, with poor prognosis and survival rates. Therefore, novel molecular targets and therapeutic strategies are needed to prolong patient survival. For this purpose, we investigated the effects of rottlerin and genistein separately and in combination on neuroblastoma cells (SH-SY5Y, Kelly). First, the effects of rottlerin and genistein were investigated on cell proliferation. Different rottlerin (1-50 µM) and genistein (5-150 µM) doses were used as experimental groups compared to the control (DMSO/vehicle). The IC50 dose was found to be 5 µM for rottlerin and 30 µM for genistein (P < 0.0001). Other analyses, such as colony formation assays, annexin V/propidium iodide staining, matrigel invasion assays, and Western blot analysis, were performed with these doses and their combinations. To assess statistical significance, statistical analysis was conducted using the one-way ANOVA with the post hoc Tukey test. Our results showed that IC50 doses of rottlerin and genistein induced a significant reduction in cell proliferation, colony formation, and invasion in neuroblastoma cells (P < 0.0001). The combination of these doses increased the levels of inhibition of cell proliferation and invasion while decreasing the level of apoptosis (P 0.0001). Furthermore, these agents caused G1-cell cycle arrest in these cells. Our western blot data showed that rottlerin and genistein treatments markedly inhibit elongation factor 2 kinase (EF2K) and other pro-tumorigenic, metastatic proteins in neuroblastoma cells. These agents probably showed their anti-proliferative, anti-metastatic, and pro-apoptotic effects through EF2K downregulation. Our results suggested that rottlerin and genistein have inhibitory effects on cancer cell proliferation, invasion, and cell cycle and induce apoptosis in both cell lines. Combined treatment with rottlerin and genistein may be a viable approach and beneficial to neuroblastoma patients as the combined effect significantly suppresses the above-mentioned pathways.

Assessment of Prognostic Factors and Validity of Scoring Models in Childhood Autoimmune Encephalitis.

OBJECTIVE: The aim of this study is to evaluate the prognostic factors in a single-center pediatric cohort with autoimmune encephalitis. MATERIALS AND METHODS: The study group consisted of 23 pediatric autoimmune encephalitis patients (seropositive autoimmune encephalitis: 15, seronegative autoimmune encephalitis: 8). Five group prognostic parameters were evaluated: clinical manifestations, elect roenc ephal ograp hy features, magnetic resonance imaging characteristics, biomarkers, and treatment modalities. Three scoring models were applied: the Antibody Prevalence in Epilepsy and Response to Immunotherapy in Epilepsy for predicting autoimmune-related epilepsy in the whole cohort and the anti-N-methyl-d-aspartate receptor Encephalitis 1-Year Functional Status score for overall outcome in patients with anti-N-methyl-d-aspartate receptor encephalitis. RESULTS: The initial clinical spectrum of the disease was similar in the seronegative and seropositive groups. Almost half of the patients (48%) recovered without any complications with first-line immunotherapy. The patients with movement disorders in the acute phase of the disease needed more likely second-line immunotherapy (P = .039). The presence of status epilepticus at admission was significantly associated with adverse outcomes and the development of autoimmune-related epilepsy (P = .019). Autoimmune-related epilepsy was defined in an equal proportion of patients (91.5%) with 2 immune epilepsy scores (Antibody Prevalence in Epilepsy and Response to Immunotherapy in Epilepsy). The N-methyl-d-aspartate receptor Encephalitis 1-Year Functional Status score and the modified Rankin score assessed for the first-year prognosis were strongly correlated among the patients with anti-N-methyl-d-aspartate receptor encephalitis (P = .03, Spearmen's rho = 0.751). CONCLUSIONS: The presence of status epilepticus was the most important prognostic factor in the patients with the adverse outcome. The studied scoring models (Anti-N-methyl-d-aspartate receptor Encephalitis 1-Year Functional Status, Antibody Prevalence in Epilepsy, and Response to Immunotherapy in Epilepsy) have also been proven to be applicable to the pediatric age group for predicting overall outcome and autoimmune-related epilepsy.

Inhibition of Sphingosine-1-Phosphate Receptor 2 by JTE013 Enhanced Alveolar Bone Regeneration by Promoting Angiogenesis.

Sphingosine-1-phosphate receptor 2 (S1PR2) is a G protein-coupled receptor that regulates various immune responses. Herein, we report the effects of a S1PR2 antagonist (JTE013) on bone regeneration. Murine bone marrow stromal cells (BMSCs) were treated with dimethylsulfoxide (DMSO) or JTE013 with or without infection by an oral bacterial pathogen Aggregatibacter actinomycetemcomitans. Treatment with JTE013 enhanced vascular endothelial growth factor A (VEGFA), platelet derived growth factor subunit A (PDGFA), and growth differentiation factor 15 (GDF15) gene expression and increased transforming growth factor beta (TGFß)/Smad and Akt signaling. Eight-week-old male C57BL/6J mice were challenged with ligatures around the left maxillary 2nd molar for 15 days to induce inflammatory bone loss. After ligature removal, mice were treated with diluted DMSO or JTE013 in the periodontal tissues 3 times per week for 3 weeks. Calcein was also injected twice to measure bone regeneration. Micro-CT scanning of maxillary bone tissues and calcein imaging revealed that treatment with JTE013 enhanced alveolar bone regeneration. JTE013 also increased VEGFA, PDGFA, osteocalcin, and osterix gene expressions in the periodontal tissues compared to control. Histological examination of periodontal tissues revealed that JTE013 promoted angiogenesis in the periodontal tissues compared to control. Our findings support that inhibition of S1PR2 by JTE013 increased TGFß/Smad and Akt signaling; enhanced VEGFA, PDGFA, and GDF15 gene expression; and subsequently promoted angiogenesis and alveolar bone regeneration.

Shared Biological Pathways and Processes in Patients with Intellectual Disability: A Multicenter Study.

BACKGROUND: Although the underlying genetic causes of intellectual disability (ID) continue to be rapidly identified, the biological pathways and processes that could be targets for a potential molecular therapy are not yet known. This study aimed to identify ID-related shared pathways and processes utilizing enrichment analyses. METHODS: In this multicenter study, causative genes of patients with ID were used as input for Disease Ontology (DO), Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes enrichment analysis. RESULTS: Genetic test results of 720 patients from 27 centers were obtained. Patients with chromosomal deletion/duplication, non-ID genes, novel genes, and results with changes in more than one gene were excluded. A total of 558 patients with 341 different causative genes were included in the study. Pathway-based enrichment analysis of the ID-related genes via ClusterProfiler revealed 18 shared pathways, with lysine degradation and nicotine addiction being the most common. The most common of the 25 overrepresented DO terms was ID. The most frequently overrepresented GO biological process, cellular component, and molecular function terms were regulation of membrane potential, ion channel complex, and voltage-gated ion channel activity/voltage-gated channel activity, respectively. CONCLUSION: Lysine degradation, nicotine addiction, and thyroid hormone signaling pathways are well-suited to be research areas for the discovery of new targeted therapies in ID patients.

The importance microRNAs as a biomarker in lung cancer.

INTRODUCTION: Lung cancer (LC) is the most common cancer  in the world.Well known  causes are  long term  smoking, environmental influences and genetic variations. LC  is divided into two main types based on their histological phenotypes; small cell lung cancer (SCLC), and non-small cell lung cancer (NSCLC). The high specificity of these new screening methods, which are non-invasive, safe, inexpensive and simple to perform, is important in the early diagnosis and prognosis of cancer. MicroRNAs are  significant biomarkers on the diagnosis metastasis and targeted therapies of NSCLC. In our study, we aimed to investigate the potential of using microRNAs as a biomarker in the early diagnosis of lung cancer. MATERIAL AND METHOD: Twenty patients diagnosed with lung cancer and  twenty healthy individuals of the same age and gender were selected as the control group.  Sixteen microRNAs were studied from blood samples. RESULT: Sixteen miRNAs (Let -7c, Let-7g, miR-1, miR-21, miR-29a, miR-31, miR-34a, miR 103a, miR-141, miR-155, miR-193b, miR-200b, miR-205, miR-340, miR-486, miR-708) were selected for tests and MiR 181 and miR 192 were used as the endogenous control group in line with their binding potentials and gene expression levels. The most specific and sensitive miRNAs were mirR-29a, miR-103a, and miR486 according to endogen controls in patients and healthy subjects. DISCUSSION: A meta-analysis study showed that circulating miRNAs could be promising biomarkers for early diagnosis of lung cancer. Overall, 17 studies were included evaluating 35 miRNA markers and 19 miRNA panels in serum or plasma. The potential role of circulating miRNAs for non-invasive lung screening has been highlighted. In conclusion, there is a need for further validation studies for the use of three  miRNAs as a biomarker in the early diagnosis and prognosis of lung cancer.