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Purpose:Using next-generation sequencing (NGS), we recently documented T-cell oligoclonality in treatment-naïve chronic lymphocytic leukemia (CLL), with evidence indicating T-cell selection by restricted antigens.Experimental Design:Here, we sought to comprehensively assess T-cell repertoire changes during treatment in relation to (i) treatment type [fludarabine-cyclophosphamide-rituximab (FCR) versus ibrutinib (IB) versus rituximab-idelalisib (R-ID)], and (ii) clinical response, by combining NGS immunoprofiling, flow cytometry, and functional bioassays.Results:T-cell clonality significantly increased at (i) 3 months in the FCR and R-ID treatment groups, and (ii) over deepening clinical response in the R-ID group, with a similar trend detected in the IB group. Notably, in constrast to FCR that induced T-cell repertoire reconstitution, B-cell receptor signaling inhibitors (BcRi) preserved pretreatment clones. Extensive comparisons both within CLL as well as against T-cell receptor sequence databases showed little similarity with other entities, but instead revealed major clonotypes shared exclusively by patients with CLL, alluding to selection by conserved CLL-associated antigens. We then evaluated the functional effect of treatments on T cells and found that (i) R-ID upregulated the expression of activation markers in effector memory T cells, and (ii) both BcRi improved antitumor T-cell immune synapse formation, in marked contrast to FCR.Conclusions:Taken together, our NGS immunoprofiling data suggest that BcRi retain T-cell clones that may have developed against CLL-associated antigens. Phenotypic and immune synapse bioassays support a concurrent restoration of functionality, mostly evident for R-ID, arguably contributing to clinical response.

Supplementary Data

Although non-surgical periodontal treatment is considered the gold standard, a subgroup of patients displays recurrence/progression of periodontitis after treatment. The aim of the present prospective study was to assess the effect of IL-6 -572 G/C and IL-10 -592 C/A gene polymorphisms on the risk of disease recurrence/progression at 3 years following non-surgical periodontal treatment.Thirty-seven patients diagnosed with chronic periodontitis received oral hygiene instructions and nonsurgical periodontal treatment and were monitored for 3 years. All individuals were clinically evaluated for PPD, CAL and BOP at baseline and 3 years. Based on the clinical findings at 3 years, all subjects were considered either "at risk" or "not at risk" of periodontal disease progression based on specific criteria. Blood samples were collected at baseline and genotyping of the polymorphisms in IL-6 (rs1800796) and IL-10 (rs1800872) genes were performed by PCR.Following DNA separation and genotyping, 70.3% of the patients were homozygous carriers of the IL-6 - 572G and 45.9% were carriers of the IL-10 -592A allele. Individuals at risk of disease progression ranged from 16.2% to 56.8% based on the criteria used. IL-6 -572 G/C and IL-10 -592 C/A polymorphisms were not associated with an increased risk of further disease progression (p0.05) when the three criteria were examined. All examined periodontal clinical measures were significantly improved (p0.05) after treatment. Males showed a significantly higher risk of disease progression than females when full-mouth BOP≥30% was considered (p=0.008).Within the limitations of this 3-year prospective study, individuals susceptible to periodontal disease as determined by the presence of the IL-6 -572GG genotype or the IL-10 -592A allele were not associated with an increased risk of further disease progression and the potential need for further treatment following non-surgical periodontal treatment. Males were more prone to be at risk of disease progression than females.

The TA-isoform of the p63 transcription factor (TAp63) has been reported to contribute to clinical aggressiveness in chronic lymphocytic leukemia (CLL) in a hitherto elusive way. Here, we sought to further understand and define the role of TAp63 in the pathophysiology of CLL. First, we found that elevated TAp63 expression levels are linked with adverse clinical outcomes, including disease relapse and shorter time-to-first treatment and overall survival. Next, prompted by the fact that TAp63 participates in an NF-κB/TAp63/BCL2 antiapoptotic axis in activated mature, normal B cells, we explored molecular links between TAp63 and BCL2 also in CLL. We documented a strong correlation at both the protein and the messenger RNA (mRNA) levels, alluding to the potential prosurvival role of TAp63. This claim was supported by inducible downregulation of TAp63 expression in the MEC1 CLL cell line using clustered regularly interspaced short palindromic repeats (CRISPR) system, which resulted in downregulation of BCL2 expression. Next, using chromatin immunoprecipitation (ChIP) sequencing, we examined whether BCL2 might constitute a transcriptional target of TAp63 and identified a significant binding profile of TAp63 in the BCL2 gene locus, across a genomic region previously characterized as a super enhancer in CLL. Moreover, we identified high-confidence TAp63 binding regions in genes mainly implicated in immune response and DNA-damage procedures. Finally, we found that upregulated TAp63 expression levels render CLL cells less responsive to apoptosis induction with the BCL2 inhibitor venetoclax. On these grounds, TAp63 appears to act as a positive modulator of BCL2, hence contributing to the antiapoptotic phenotype that underlies clinical aggressiveness and treatment resistance in CLL.

Purpose: Using next-generation sequencing (NGS), we recently documented T-cell oligoclonality in treatment-naïve chronic lymphocytic leukemia (CLL), with evidence indicating T-cell selection by restricted antigens.Experimental Design: Here, we sought to comprehensively assess T-cell repertoire changes during treatment in relation to (i) treatment type [fludarabine-cyclophosphamide-rituximab (FCR) versus ibrutinib (IB) versus rituximab-idelalisib (R-ID)], and (ii) clinical response, by combining NGS immunoprofiling, flow cytometry, and functional bioassays.Results: T-cell clonality significantly increased at (i) 3 months in the FCR and R-ID treatment groups, and (ii) over deepening clinical response in the R-ID group, with a similar trend detected in the IB group. Notably, in constrast to FCR that induced T-cell repertoire reconstitution, B-cell receptor signaling inhibitors (BcRi) preserved pretreatment clones. Extensive comparisons both within CLL as well as against T-cell receptor sequence databases showed little similarity with other entities, but instead revealed major clonotypes shared exclusively by patients with CLL, alluding to selection by conserved CLL-associated antigens. We then evaluated the functional effect of treatments on T cells and found that (i) R-ID upregulated the expression of activation markers in effector memory T cells, and (ii) both BcRi improved antitumor T-cell immune synapse formation, in marked contrast to FCR.Conclusions: Taken together, our NGS immunoprofiling data suggest that BcRi retain T-cell clones that may have developed against CLL-associated antigens. Phenotypic and immune synapse bioassays support a concurrent restoration of functionality, mostly evident for R-ID, arguably contributing to clinical response.

Background: Oxidation reactions are known to shorten the shelf life and cause damage to foods rich in fat, such as dairy products. One way to limit oxidation and increase the shelf life of fermented dairy products is to use natural antioxidants. The aim of this study was to examine the effect of adding pomegranate extracts in the antioxidant properties, rheological characteristics and the storage stability of the fermented product of kefir. Methods: The Pomegranate Juice (PGJ) and Peel Extracts (PGPE) (5%, 10% w/v) were added to kefir and the antioxidant properties were evaluated by using the methods of radical scavenging activity (DPPH) and Ferric Reducing Antioxidant Power Activity (FRAP). Spectrophotometric and instrumental methods were used to determine the Total Phenols (TPs), pH values, viscosity and flow behavioral index values of enriched with pomegranate kefir samples. The same properties were tested when kefir samples stored at 4°C for 7, 14, 21 and 28 days. Results: The addition of PGJ and PGPE results in an increase in the antioxidant activity (DPPH, FRAP) and total phenol content (TPs) of kefir samples. Increasing the concentration of the added PGJ and PGPE, results in an increase in the TP content and the DPPH activity of kefir. As far as the storage time is concerned, the results showed an increase in the amount of TP at 7th day and a reduction in the DPPH activity in the 14th day of storage. In contrary to the DPPH method, the increase in storage time has resulted in a reduction in antioxidant activity by the FRAP method. The addition of PGJ and PGPE in kefir results in a decrease in pH values while the pH of kefir samples increased during storage at 4°C for 28 days. The addition of PGJ and PGPE to kefir samples results to a decrease in viscosity and an increase in the flow behavior index. Increasing storage time results in increased flow behavior index of kefir samples. Conclusion: The addition of PGJ and PGPE increased the antioxidant activity and total phenols of the kefir product and preserved its properties during the total storage time of 28 days at 4°C.

The complexity of the admixture dynamics that shaped American populations is unveiled by Ongaro et al., where genetic data for more than 12,000 individuals from the continents are investigated. This study evaluates the dramatic impact of events after the colonial era, revealing a spatial and temporal heterogeneity and mirroring historical records. © 2019 Elsevier Ltd The human genetic diversity of the Americas has been affected by several events of gene flow that have continued since the colonial era and the Atlantic slave trade. Moreover, multiple waves of migration followed by local admixture occurred in the last two centuries, the impact of which has been largely unexplored. Here, we compiled a genome-wide dataset of ∼12,000 individuals from twelve American countries and ∼6,000 individuals from worldwide populations and applied haplotype-based methods to investigate how historical movements from outside the New World affected (1) the genetic structure, (2) the admixture profile, (3) the demographic history, and (4) sex-biased gene-flow dynamics of the Americas. We revealed a high degree of complexity underlying the genetic contribution of European and African populations in North and South America, from both geographic and temporal perspectives, identifying previously unreported sources related to Italy, the Middle East, and to specific regions of Africa. © 2019 Elsevier Ltd Preprint version of the article: "The genomic impact of European colonization of the Americas", posted June 28, 2019 on bioRxiv. Article is now published in Current Biology doi: [dx.doi.org/10.1016/j.cub.2019.09.076]. Published version on this repository: [http://vinar.vin.bg.ac.rs/handle/123456789/8654].

Background: Oxidation reactions are known to shorten the shelf life and cause damage to foods rich in fat, such as dairy products. One way to limit oxidation and increase the shelf life of fermented dairy products is to use natural antioxidants. The aim of this study was to examine the effect of adding pomegranate extracts in the antioxidant properties, rheological characteristics and the storage stability of the fermented product of kefir. Methods: The Pomegranate Juice (PGJ) and Peel Extracts (PGPE) (5%, 10% w/v) were added to kefir and the antioxidant properties were evaluated by using the methods of radical scavenging activity (DPPH) and Ferric Reducing Antioxidant Power Activity (FRAP). Spectrophotometric and instrumental methods were used to determine the Total Phenols (TPs), pH values, viscosity and flow behavioral index values of enriched with pomegranate kefir samples. The same properties were tested when kefir samples stored at 4°C for 7, 14, 21 and 28 days. Results: The addition of PGJ and PGPE results in an increase in the antioxidant activity (DPPH, FRAP) and total phenol content (TPs) of kefir samples. Increasing the concentration of the added PGJ and PGPE, results in an increase in the TP content and the DPPH activity of kefir. As far as the storage time is concerned, the results showed an increase in the amount of TP at 7th day and a reduction in the DPPH activity in the 14th day of storage. In contrary to the DPPH method, the increase in storage time has resulted in a reduction in antioxidant activity by the FRAP method. The addition of PGJ and PGPE in kefir results in a decrease in pH values while the pH of kefir samples increased during storage at 4°C for 28 days. The addition of PGJ and PGPE to kefir samples results to a decrease in viscosity and an increase in the flow behavior index. Increasing storage time results in increased flow behavior index of kefir samples. Conclusion: The addition of PGJ and PGPE increased the antioxidant activity and total phenols of the kefir product and preserved its properties during the total storage time of 28 days at 4°C. © 2019 Bentham Science Publishers.

The human genetic diversity of the Americas has been shaped by several events of gene flow that have continued since the Colonial Era and the Atlantic slave trade. Moreover, multiple waves of migration followed by local admixture occurred in the last two centuries, the impact of which has been largely unexplored. Here we compiled a genome-wide dataset of ~12,000 individuals from twelve American countries and ~6,000 individuals from worldwide populations and applied haplotype-based methods to investigate how historical movements from outside the New World affected i) the genetic structure, ii) the admixture profile, iii) the demographic history and iv) sex-biased gene-flow dynamics, of the Americas. We revealed a high degree of complexity underlying the genetic contribution of European and African populations in North and South America, from both geographic and temporal perspectives, identifying previously unreported sources related to Italy, the Middle East and to specific regions of Africa.

Susceptible genotypes to periodontal disease are associated with disease onset and progression. The aim of this study was to examine the effect of gene polymorphisms on the risk of further disease progression and the need for further treatment among adults with chronic periodontal disease. Sixty-seven patients diagnosed with chronic periodontitis were grouped according to genotype status and risk of further progression of disease and tooth loss. All individuals were clinically evaluated for probing pocket depth, clinical attachment loss and bleeding on probing at baseline and 45 days after treatment. Blood samples were collected at baseline and genotyping of the polymorphisms in IL-6 (rs1800796) and IL-10 (rs1800872) genes were performed by PCR. Following DNA separation and genotyping, 65.7% of the patients were homozygous carriers of the IL-6 −572G and 49.3% were carriers of the IL-10 −592A allele. Individuals at risk of disease progression ranged from 7.5% to 62.7% based on the criteria used. Carriers of the IL-10 −592A allele were significantly associated with BOP ≥ 30% and therefore exhibited a higher risk of further periodontal breakdown (p = 0.018) with an odds ratio of 1.18. None of the other definitions of disease progression were significantly associated with the examined IL-6 and IL-10 genotypes (p > 0.05). IL-10 polymorphism was associated with an increased risk of further disease progression and the potential need for further treatment following non-surgical periodontal treatment. Susceptible IL-6 genotypes were not associated with the risk of persisting or recurrent disease activity.

**Background**\ud \ud Studies investigating associations between ACTN3 R577X and ACE I/D genotypes and endurance athletic status have been limited by small sample sizes from mixed sport disciplines and lack quantitative measures of performance. Aim: To examine the association between ACTN3 R577X and ACE I/D genotypes and best personal running times in a large homogeneous cohort of endurance runners.\ud \ud **Methods**\ud \ud We collected a total of 1064 personal best 1500, 3000, 5000 m and marathon running times of 698 male and female Caucasian endurance athletes from six countries (Australia, Greece, Italy, Poland, Russia and UK). Athletes were genotyped for ACTN3 R577X and ACE ID variants.\ud \ud **Results**\ud \ud There was no association between ACTN3 R577X or ACE I/D genotype and running performance at any distance in men or women. Mean (SD) marathon times (in s) were for men: ACTN3 RR 9149 (593), RX 9221 (582), XX 9129 (582) p = 0.94; ACE DD 9182 (665), ID 9214 (549), II 9155 (492) p = 0.85; for women: ACTN3 RR 10796 (818), RX 10667 (695), XX 10675 (553) p = 0.36; ACE DD 10604 (561), ID 10766 (740), II 10771 (708) p = 0.21. Furthermore, there were no associations between these variants and running time for any distance in a sub-analysis of athletes with personal records within 20% of world records.\ud \ud **Conclusions**\ud \ud Thus, consistent with most case-control studies, this multi-cohort quantitative analysis demonstrates it is unlikely that ACTN3 XX genotype provides an advantage in competitive endurance running performance. For ACE II genotype, some prior studies show an association but others do not. Our data indicate it is also unlikely that ACE II genotype provides an advantage in endurance running.