CDX2 Research Articles

Association of 25(OH)-Vitamin D3 Serum Concentrations and Vitamin D Receptor Gene Variants with the Risk of Idiopathic Spontaneous Preterm Birth in the Croatian Population

Preterm birth (PTB) forms a heterogeneous group with possible genetic predisposition. 25(OH)-vitamin D3 plays a significant role during implantation, placentation, and the maintenance of normal pregnancy. The aim of our research was to examine whether FokI, Cdx2, and ApaI VDR gene variants, as well as serum concentrations of 25-hydroxy25(OH)-vitamin D3 in women and their newborns, might be predisposing factors for idiopathic spontaneous preterm birth. The patient group consisted of 44 pairs of women with ISPTB and their children, and the control group consisted of 44 pairs of women who delivered at term and their children. At the time of delivery, peripheral blood was collected from every woman, and after newborn delivery, umbilical cord blood was collected. For genotyping of the rs2228570 C/T, rs11568820 G/A, and rs7975232 T/G SNPs, a combination of polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) was used. Serum concentrations of 25(OH)-vitamin D3 were determined using high-performance liquid chromatography (HPLC). There were no statistically significant differences in the frequencies of VDR genotypes and alleles between women with ISPTB and control women. There was a statistically significant difference in the distribution of VDR Cdx-2 (rs11568820) genotypes between preterm-born children and controls, with the GG genotype and G allele being more prevalent among patients than controls (p < 0.001). There were no statistically significant differences in mean values between women with ISPTB and control women, nor between preterm and full-term newborns, although the 25(OH)-vitamin D3 concentrations in preterm-born children were lower than in controls. Furthermore, there was a statistically significant correlation in 25(OH)-vitamin D3 concentrations between mothers and children both in the patient and in the control groups (b = 0.771, p < 0.001). The results of our study demonstrate a notable association between the VDR Cdx2 gene polymorphism and idiopathic spontaneous preterm birth (ISPTB) in a Caucasian population, but because of the small number of participants, further research is needed.

The Effect of Simulated Physiological Oocyte Maturation (SPOM) and L-Carnitine on Bovine Oocyte Developmental Competence

Background: Simulated Physiological Oocyte Maturation (SPOM) mimics in vitro the physiological events of oocyte maturation in the presence of cAMP modulators. These modulators increase the intracellular concentrations of cAMP, which inhibits the immediate resumption of meiosis and gives the oocyte more time to gain optimal developmental competence. In addition, L-carnitine helps to increase the energy supply of cells through the β-oxidation of fatty acids. This study aimed to investigate the effect of SPOM and L-carnitine supplementation during In Vitro Maturation (IVM) and In Vitro Culture (IVC) on the developmental competence of bovine oocytes. Methods: Ovarian Cumulus Complexes (COCs) were cultured in the presence or absence of forskolin+IBMX during the first 2 hr of IVM (pre-IVM) with or without L-carnitine (LC) during IVM or IVC in six experimental groups as follows: I) pre-IVM (pre-IVM group), II) pre-IVM with L-carnitine supplementation during IVM (pre-IVM/LC group), III) L-carnitine supplementation during IVM (IVM/LC group), IV) L-carnitine supplementation during in vitro culture (IVC/LC group), V) pre-IVM+ IVC/LC group, and VI) no treatment during IVM and IVC (Control group). The cleavage and blastocyst rates, the blastocysts’ total cells number, and the expression of Nanog, Bax, Oct4, Cdx2, and Ifnt genes in resulting blastocysts were assessed. To assess differences among experimental groups, a one-way analysis of variance was initially employed, followed by post hoc Fisher LSD. The difference between groups was considered statistically significant when p<0.05. Results: The cleavage and blastocyst rates in the Pre-IVM and Pre-IVM/LC groups was higher than control group and other groups (p≤0.05) except for IVC/LC and IVM/LC groups, respectively. The number of blastocyst’s Inner Cell Mass (ICM) in pre-IVM and Pre-IVM/LC groups as well as the ratio of ICM/TE were higher than control group (p<0.05). The expression of OCT4, CDX2, and IFNT increased in both the pre-IVM and pre-IVM/LC groups compared to the control group (p<0.05). Conclusion: In conclusion, the application of SPOM-adapted IVM and L-carnitine during IVM of bovine oocyte improves the quantity and quality of the resulting embryos.

Vitamin C Improves Oocyte In Vitro Maturation and Potentially Changes Embryo Quality in Cattle.

To obtain high-quality bovine oocytes, the effects of vitamin C (VC) on the IVM of bovine oocytes and early embryo development were investigated. The results showed the following. (1) The IVM medium containing 50 µg/mL VC improved the oocyte maturation rate but did not affect the parthenogenetic embryo development. (2) The IVC medium containing 20 µg/mL VC improved the cleavage rate of the IVF embryos and enhanced the mRNA transcriptions of pluripotency gene Oct4, Sox2, Cdx2, and Nanog in the blastocysts but had no effects on the blastocyst rate. (3) Combining supplementation of 50 µg/mL VC in IVM medium + 20 µg/mL VC in IVC medium (named as VC 50/20, similar hereinafter) elevated the cleavage rate of IVF embryos and enhanced the mRNA expressions of Oct4, Sox2, Cdx2, and Nanog in the blastocysts. (4) Combination of VC 0/20 and VC 50/20 enhanced the transcription of anti-apoptotic gene Bcl-2 and VC 50/0 weakened the transcription of pro-apoptotic gene Bax, while VC 0/40 and VC 0/60 increased Bax expression and diminished the Bcl-2/Bax ratio in blastocysts. Together, employing 50 µg/mL VC improves the IVM of bovine oocytes and combination of VC 50/20 potentially changes bovine embryo quality by enhancing the expressions of the pluripotency genes and regulating the expressions of apoptosis-related genes.

In vitro developmental competence of bovine demi-embryos generated by blastomere separation and blastocyst bisection.

The efficiency of bovine invitro embryo production can be significantly improved by splitting embryos at different stages. However, the blastocyst quality of invitro-produced demi-embryos remains unexplored. The objective of this research was to compare embryo developmental rates and quality of bovine demi-embryos produced by two different strategies: (a) embryo bisection (BSEC) and (b) 2-cell blastomere separation (BSEP). To determine demi-embryos quality, we evaluated total blastocyst cell number and proportion of SOX2+ cells. Additionally, the expression of SOX2, NANOG, OCT4, CDX2, IFNT, BAX and BCL genes and let-7a and miRNA-30c Micro RNAs was analysed. BSEP resulted in improved blastocyst development, higher ICM cells and a significantly higher expression of IFNΤ than demi-embryos produced by BSEC. Let-7a, which is associated with low pregnancy establishment was detected in BSEC, while miRNA-30c expression was observed in all treatments. In conclusion, BSEP of 2-cell embryos is more efficient to improve invitro bovine embryo development and to produce good quality demi-embryos based on ICM cell number and the expression pattern of the genes explored compared to BSEC.

Upstream regulation of neurotransmitter signaling genes in metastatic colorectal cancer (mCRC) patients: Data from CALGB/SWOG 80405 trial (Alliance) and MONSTAR-SCREEN-2.

3523 Background: Evidence suggests neurotransmitter signaling may play a crucial role in tumorigenesis of gastrointestinal (GI) cancers, including migration, proliferation, and apoptosis, as well as angiogenesis and the microenvironment. We developed and applied a strategy to identify upstream regulators of neurotransmitters and their associations with patient outcomes. Methods: Prior literature yielded 85 neurotransmitter signaling genes mapped to RNA-seq gene expression in tumor tissue from 545 patients with mCRC enrolled in the phase III CALGB/SWOG 80405 trial. These samples were also sequenced by FoundationOne CDx for DNA damage. Using Cox proportional hazards regression, we identified a subset of CDx genes associated with overall survival (OS) or progression-free survival (PFS). We extended the Causal Inference Test (CIT) for time-to-event data and identified a subset of the 85 signaling genes consistent with mediation of DNA mutation effects on OS and PFS. Another CIT analysis identified potential upstream neurotransmitter regulators, gene expression mediators of mutation effects on the signaling genes. Top genes were then assessed in whole-transcriptome sequencing (WTS) expression data with Caris MI profile collected from 398 patients with advanced CRC enrolled in the MONSTAR-SCREEN-2 trial. Results: At the 0.05 alpha level, we identified 20 signaling genes with gene expression associated with PFS and/or OS, and 68 genes with evidence of DNA mutations associated with PFS and/or OS. False discovery rate (FDR) < 0.05 was applied to filter the 56,674 expression features on PFS and OS associations, yielding 173 unique genes as possible mediators. The CIT yielded 53 putative mediators at FDR < 0.05, suggesting upstream regulation of neurotransmitter signaling. Five genes were identified as targeting at least 8 signaling genes each, PIGU, ANO9, LY6G6D, and GPR160, and FSCN1, where the first 4 are favorable and only FSCN1 is a poorer prognostic. Following up on these 5 genes in MONSTAR-SCREEN-2 by dichotomizing at the median value of WTS expression, 3 were nominally associated with OS, PIGU ( P = 0.050; HR, 0.59; 95%CI, 0.34-1.00), ANO9( P = 0.029; HR, 0.55; 95%CI, 0.32-0.95), LY6G6D( P = 0.0024; HR, 0.44; 95%CI, 0.25-0.76), and FSCN1 was suggestive ( P = 0.079; HR, 1.62; 95%CI, 0.94-2.77), with direction of effects in agreement with 80405 results for all four. Conclusions: Our novel approach identified previously unknown upstream regulators of neurotransmitter signaling associated with treatment outcomes in mCRC. PIGU, FSCN1, and LY6G6D have been suggested in prior literature as targets for CRC and ANO9 has been suggested as a target for GI cancer. These results bolster the strategy of using mediation analysis with large-scale molecular data to identify new therapeutic targets in CRC. NCT00265850.

Evaluation of the Immunohistochemical Scoring System of CDX2 Expression as a Prognostic Biomarker in Colon Cancer.

Encoded by the CDX2 homeobox gene, the CDX2 protein assumes the role of a pivotal transcription factor localized within the nucleus of intestinal epithelial cells, orchestrating the delicate equilibrium of intestinal physiology while intricately guiding the precise development and differentiation of epithelial tissue. Emerging research has unveiled that positive immunohistochemical expression of this protein shows that the CDX2 gene exerts a potent suppressive impact on tumor advancement in colorectal cancer, impeding the proliferation and distant dissemination of tumor cells, while the inhibition or suppression of CDX2 frequently correlates with aggressive behavior in colorectal cancer. In this study, we conducted an immunohistochemical assessment of CDX2 expression on a cohort of 43 intraoperatively obtained tumor specimens from patients diagnosed with colon cancer at Colțea Clinical Hospital in Bucharest, between April 2019 and December 2023. Additionally, we shed light on the morphological diversity within colon tumors, uncovering varying differentiation grades within the same tumor, reflecting the variations in CDX2 expression as well as the genetic complexity underlying these tumors. Based on the findings, we developed an innovative immunohistochemical scoring system that addresses the heterogeneous nature of colon tumors. Comprehensive statistical analysis of CDX2 immunohistochemical expression unveiled significant correlations with known histopathological parameters such as tumor differentiation grades (p-value = 0.011) and tumor budding score (p-value = 0.002), providing intriguing insights into the complex involvement of the CDX2 gene in orchestrating tumor progression through modulation of differentiation processes, and highlighting its role in metastatic predisposition. The compelling correlation identified between CDX2 expression and conventional histopathological parameters emphasizes the prognostic significance of the CDX2 biomarker in colon cancer. Moreover, our novel immunohistochemical scoring system reveals a distinct subset of colon tumors exhibiting reserved prognostic outcomes, distinguished by their "mosaic" CDX2 expression pattern.

Deubiquitylase ubiquitin-specific protease 7 plays a crucial role in the lineage differentiation of preimplantation blastocysts†.

Preimplantation embryos undergo a series of important biological events, including epigenetic reprogramming and lineage differentiation, and the key genes and specific mechanisms that regulate these events are critical to reproductive success. Ubiquitin-specific protease 7 (USP7) is a deubiquitinase involved in the regulation of a variety of cellular functions, yet its precise function and mechanism in preimplantation embryonic development remain unknown. Our results showed that RNAi-mediated silencing of USP7 in mouse embryos or treatment with P5091, a small molecule inhibitor of USP7, significantly reduced blastocyst rate and blastocyst quality, and decreased total and trophectoderm cell numbers per blastocyst, as well as destroyed normal lineage differentiation. The results of single-cell RNA-seq, reverse transcription-quantitative polymerase chain reaction, western blot, and immunofluorescence staining indicated that interference with USP7 caused failure of the morula-to-blastocyst transition and was accompanied by abnormal expression of key genes (Cdx2, Oct4, Nanog, Sox2) for lineage differentiation, decreased transcript levels, increased global DNA methylation, elevated repressive histone marks (H3K27me3), and decreased active histone marks (H3K4me3 and H3K27ac). Notably, USP7 may regulate the transition from the morula to blastocyst by stabilizing the target protein YAP through the ubiquitin-proteasome pathway. In conclusion, our results suggest that USP7 may play a crucial role in preimplantation embryonic development by regulating lineage differentiation and key epigenetic modifications.

A patterned human neural tube model using microfluidic gradients.

The human nervous system is a highly complex but organized organ. The foundation of its complexity and organization is laid down during regional patterning of the neural tube, the embryonic precursor to the human nervous system. Historically, studies of neural tube patterning have relied on animal models to uncover underlying principles. Recently, models of neurodevelopment based on human pluripotent stem cells, including neural organoids1-5 and bioengineered neural tube development models6-10, have emerged. However, such models fail to recapitulate neural patterning along both rostral-caudal and dorsal-ventral axes in a three-dimensional tubular geometry, a hallmark of neural tube development. Here we report a human pluripotent stem cell-based, microfluidic neural tube-like structure, the development of which recapitulates several crucial aspects of neural patterning in brain and spinal cord regions and along rostral-caudal and dorsal-ventral axes. This structure was utilized for studying neuronal lineage development, which revealed pre-patterning of axial identities of neural crest progenitors and functional roles of neuromesodermal progenitors and the caudal gene CDX2 in spinal cord and trunk neural crest development. We further developed dorsal-ventral patterned microfluidic forebrain-like structures with spatially segregated dorsal and ventral regions and layered apicobasal cellular organizations that mimic development of the human forebrain pallium and subpallium, respectively. Together, these microfluidics-based neurodevelopmentmodels provide three-dimensional lumenal tissue architectures with in vivo-like spatiotemporal cell differentiation and organization, which will facilitate the study of human neurodevelopment and disease.

TEAD4 regulates KRT8 and YAP in preimplantation embryos in mice but not in cattle.

Lineage specification plays a vital role in preimplantation development. TEAD4 is an essential transcription factor for trophectoderm lineage specification in mice but not in cattle. Tead4, a critical transcription factor expressed during preimplantation development, is essential for the expression of trophectoderm-specific genes in mice. However, the functional mechanism of TEAD4 in mouse preimplantation development and its conservation across mammals remain unclear. Here, we report that Tead4 is a crucial transcription factor necessary for blastocyst formation in mice. Disruption of Tead4 through base editing results in developmental arrest at the morula stage. Additionally, RNA-seq analysis reveals dysregulation of 670 genes in Tead4 knockout embryos. As anticipated, Tead4 knockout led to a decrease in trophectoderm genes Cdx2 and Gata3. Intriguingly, we observed a reduction in Krt8, suggesting that Tead4 influences the integrity of the trophectoderm epithelium in mice. More importantly, we noted a dramatic decrease in nuclear Yap in outside cells for Tead4-deficient morula, indicating that Tead4 directly regulates Hippo signaling. In contrast, bovine embryos with TEAD4 depletion could still develop to blastocysts with normal expression of CDX2, GATA3, and SOX2, albeit with a decrease in total cell number and ICM cell number. In conclusion, we propose that Tead4 regulates mouse blastocyst formation via Krt8 and Yap, both of which are critical regulators of mouse preimplantation development.

Expression and Promoter Methylation Status of CDX1 and CDX2 Genes in Chinese Patients with Gastric Cancer

Expression and Promoter Methylation Status of CDX1 and CDX2 Genes in Chinese Patients with Gastric Cancer

CDX1 Gene Expression in Recurrent Spontaneous Abortion under Extracellular Regulatory Protein Kinase1/2 Signaling Pathway

CDX1 Gene Expression in Recurrent Spontaneous Abortion under Extracellular Regulatory Protein Kinase1/2 Signaling Pathway

Human RFX6 regulates endoderm patterning at the primitive gut tube stage.

Transcriptional factor RFX6 is known to be a causal gene of Mitchell-Riley syndrome (MRS), an autosomal recessive neonatal diabetes associated with pancreatic hypoplasia and intestinal atresia/malformation. The morphological defects are limited to posterior foregut and mid-hindgut endodermal lineages and do not occur in the anterior foregut lineage; the mechanism remains to be fully elucidated. In this study, we generated RFX6+/eGFP heterozygous knockin and RFX6eGFP/eGFP homozygous knockin/knockout human-induced pluripotent stem cell (hiPSC) lines and performed in vitro endoderm differentiation to clarify the role of RFX6 in early endoderm development. RFX6 expression was found to surge at the primitive gut tube (PGT) stage in comparison with that in the undifferentiated or definitive endoderm stage. At the PGT stage, the expression of PDX1 and CDX2, posterior foregut and mid-hindgut master regulators, respectively, was decreased by the RFX6 deficit. PDX1+ and CDX2+ cells were mostly green fluorescent protein (GFP)+ in RFX6+/eGFP hiPSCs, but their cell number was markedly decreased in RFX6eGFP/eGFP hiPSCs. The expression of SOX2, an anterior foregut marker, was not affected by the RFX6 deficit. In addition, we found a putative RFX6-binding X-box motif using cap analysis of gene expression-seq and the motif-containing sequences in the enhancer regions of PDX1 and CDX2 bound to RFX6 in vitro. Thus, RFX6 regulates the ParaHox genes PDX1 and CDX2 but does not affect SOX2 in early endodermal differentiation, suggesting that defects in early stage endoderm patterning account for the morphological pathology of MRS.

Loss of function of ribosomal protein L13a blocks blastocyst formation and reveals a potential nuclear role in gene expression.

Ribosomal proteins play diverse roles in development and disease. Most ribosomal proteins have canonical roles in protein synthesis, while some exhibit extra-ribosomal functions. Previous studies in our laboratory revealed that ribosomal protein L13a (RPL13a) is involved in the translational silencing of a cohort of inflammatory proteins in myeloid cells. This prompted us to investigate the role of RPL13a in embryonic development. Here we report that RPL13a is required for early development in mice. Crosses between Rpl13a+/- mice resulted in no Rpl13a-/- offspring. Closer examination revealed that Rpl13a-/- embryos were arrested at the morula stage during preimplantation development. RNA sequencing analysis of Rpl13a-/- morulae revealed widespread alterations in gene expression, including but not limited to several genes encoding proteins involved in the inflammatory response, embryogenesis, oocyte maturation, stemness, and pluripotency. Exvivo analysis revealed that RPL13a was localized to the cytoplasm and nucleus between the two-cell and morula stages. RNAi-mediated depletion of RPL13a phenocopied Rpl13a-/- embryos and knockdown embryos exhibited increased expression of IL-7 and IL-17 and decreased expression of the lineage specifier genes Sox2, Pou5f1, and Cdx2. Lastly, a protein-protein interaction assay revealed that RPL13a is associated with chromatin, suggesting an extra ribosomal function in transcription. In summary, our data demonstrate that RPL13a is essential for the completion of preimplantation embryo development. The mechanistic basis of the absence of RPL13a-mediated embryonic lethality will be addressed in the future through follow-up studies on ribosome biogenesis, global protein synthesis, and identification of RPL13a target genes using chromatin immunoprecipitation and RNA-immunoprecipitation-based sequencing.

Using extended growth of cattle embryos in culture to gain insights into bovine developmental events on embryonic days 8 to 10

We have previously described an extended embryo culture system, based on uterine composition, growth factors and the cell culture additive B27, for growing cattle embryos in vitro beyond embryonic day 7. Here, extended in vitro embryos are compared to embryos developed in the uterus and are used to establish a developmental staging framework useful for understanding developmental events occurring until Day 10. Immunofluorescence or mRNA expression of the ICM/epiblast markers OCT4, SOX2 and NANOG, hypoblast markers GATA6, SOX17 and GATA4 and trophoblast genes CDX2, GATA3, ASCL2 and IFNT revealed the presence of four stages during this period that can be molecularly distinguished. These are expanded blastocyst, hatched blastocyst, hypoblast layering and early hypoblast migration. Interestingly NANOG and SOX17 show reciprocal expression at the expanded blastocyst stage, well before SOX2 and GATA6 expression refines to a similar so-called “salt and pepper” mutually exclusive expression in the ICM at the hatched blastocyst stage. GATA4 expression is only seen from stages when the hypoblast starts migrating around the blastocyst cavity. Intriguingly, trophoblast still expresses GATA6 and OCT4 in all cells during the expanded blastocyst phase, while SOX2 and SOX17 are seen in only some trophoblast cells. By the hypoblast-epiblast layering stage no trophoblast expression remains except for that of OCT4 protein, which starts waning in trophoblast once the hypoblast begins migrating. Lastly, it is shown that cultured embryos exhibit increased expression of the stress marker TP53 in the epiblast and hypoblast at late stages in comparison to embryos produced in the uterine environment.

The Effects of Retinoic Acid on the Expression of HNF4α and Cdx-2 Genes in Caco-2 Cells as a Model of Intestinal Barrier

The Effects of Retinoic Acid on the Expression of HNF4α and Cdx-2 Genes in Caco-2 Cells as a Model of Intestinal Barrier

N4-acetylcytidine of Nop2 mRNA is required for the transition of morula-to-blastocyst.

N-acetyltransferase 10 (NAT10)-mediated N4-acetylcytidine (ac4C) modification is crucial for mRNA stability and translation efficiency, yet the underlying function in mammalian preimplantation embryos remains unclear. Here, we characterized the ac4C modification landscape in mouse early embryos and found that the majority of embryos deficient in ac4C writer-NAT10 failed to develop into normal blastocysts. Through single-cell sequencing, RNA-seq, acetylated RNA immunoprecipitation combined with PCR (acRIP-PCR), and embryonic phenotype monitoring, Nop2 was screened as a target gene of Nat10. Mechanistically, Nat10 knockdown decreases the ac4C modification on Nop2 mRNA and reduces RNA and protein abundance by affecting the mRNA stability of Nop2. Then, depletion of NOP2 may inhibit the translation of transcription factor TEAD4, resulting in defective expression of the downstream lineage-specific gene Cdx2, and ultimately preventing blastomeres from undergoing the trophectoderm (TE) fate. However, exogenous Nop2 mRNA partially reverses this abnormal development. In conclusion, our findings demonstrate that defective ac4C modification of Nop2 mRNA hinders the morula-to-blastocyst transition by influencing the first cell fate decision in mice.

New tetradentate long chain Schiff base and its cadmium(II) complexes: Antimicrobial, antioxidant and anticancer activities

In the current study, a new long chain tetra-nitrogen donor Schiff base ligand (SB) entitled as N1,N2-bis(2-(((1E,2E)-3-(4-(dimethylamino)phenyl)allylidene)amino)ethyl)ethane-1,2-diamine and some its molecular cadmium (II) complexes formulated as Cd(SB)X2 (X = halide/pseudohalide) were prepared and characterized by various physicochemical and spectral methods such as melting point, micro-analytical data, thermal analysis (TG/DTG/DTA), molar conductivity and UV–Visible, FT-IR, 1H, and 13C NMR spectroscopies. Also nanostructured cadmium chloride and iodide chelate complexes approved by PXRD SEM and EDX techniques were synthesized to use as precursor to prepare nanostructure cadmium oxide. The particle sizes about 2–10 nm were estimated for nanostructured cadmium complexes and oxide. Thermo-gravimetric plots showed the compounds are decomposed via 2–4 steps with weight loss of 69–100% to leave out metallic Cd or cadmium salt as final residue. Thermo-kinetic activation parameters calculated based on TG plots suggested non-spontaneous thermal decomposition with activation Ea*, ΔH* and ΔG* parameters in the ranges of (13.54–160.48 kJ.mol−1), (8.22–155.95 kJ.mol−1) and (1.59 × 102-4.1 × 102 kJ.mol−1) respectively. Moreover, the pharmacological activities of the compounds were investigated against some pathogenic fungi and bacteria. According to screening data, the SB ligand showed lower antimicrobial activities than its complexes toward selected microbes so that the cadmium(II) nitrate, thiocyanate and azide were found the most active against B. subtilis, S. aureus, P. aeruginosa and E. coli respectively. The evaluation of DNA cleavage potency using the agarose gel electrophoresis technique suggested Cd(SB)Cl2 as the most destructive compound. Furthermore, the antioxidant assessment of the compounds as compared with the standard references by DPPH and FRAP methods proposed Cd(SB)Cl2 and Cd(SB)Br2 as the best antioxidant. In final, the cytotoxicity of the synthesized compounds was appraised toward human breast cancer (MCF-7) and prostate cancer (PC3) cell lines employing MTT and nitric oxide level assays. The results suggested Cd(SB)Br2 as more cytotoxic than other complexes against two studied cancer cell lines.

Microbiota composition and its impact on DNA methylation in colorectal cancer.

Colorectal cancer is a complex disease resulting from the interaction of genetics, epigenetics, and environmental factors. DNA methylation is frequently found in tumor suppressor genes to promote cancer development. Several factors are associated with changes in the DNA methylation pattern, and recently, the gastrointestinal microbiota could be associated with this epigenetic change. The predominant phyla in gut microbiota are Firmicutes and Bacteroidetes; however, an enrichment of Bacteroides fragilis, Fusobacterium nucleatum, and Streptococcus bovis, among others, has been reported in colorectal cancer, although the composition could be influenced by several factors, including diet, age, sex, and cancer stage. Fusobacterium nucleatum, a gram-negative anaerobic bacillus, is mainly associated with colorectal cancer patients positive for the CpG island methylator phenotype, although hypermethylation in genes such as MLH1, CDKN2A, MTSS1, RBM38, PKD1, PTPRT, and EYA4 has also been described. Moreover, Hungatella hathewayi, a gram-positive, rod-shaped bacterium, is related to hypermethylation in SOX11, THBD, SFRP2, GATA5, ESR1, EYA4, CDX2, and APC genes. The underlying epigenetic mechanism is unclear, although it could be implicated in the regulation of DNA methyltransferases, enzymes that catalyze the transfer of a methyl group on cytosine of CpG sites. Since DNA methylation is a reversible event, changes in gut microbiota could modulate the gene expression through DNA methylation and improve the colorectal cancer prognosis.

NKX3.1 immunohistochemistry and methylome profiling in mesenchymal chondrosarcoma: additional diagnostic value for a well-defined disease?

Mesenchymal chondrosarcoma (MCS) is a rare and highly aggressive tumour of soft tissue and bone that is defined by an underlying and highly specific fusion transcript involving HEY1 and NCOA2. Histologically, the tumours show a biphasic appearance consisting of an undifferentiated blue and round cell component as well as islands of highly differentiated cartilage. Particularly in core needle biopsies, the chondromatous component can be missed and the non-specific morphology and immunophenotype of the round cell component can cause diagnostic challenges. We applied NKX3.1 immunohistochemistry which was recently reported as a highly specific marker as well as methylome and copy number profiling to a set of 45 well characterised MCS cases to evaluate their potential diagnostic value. Methylome profiling revealed a highly distinct cluster for MCS. Notably, the findings were reproducible also when analysing the round cell and cartilaginous component separately. Furthermore, four outliers were identified by methylome profiling for which the diagnosis had to be revised. NKX3.1 immunohistochemistry showed positivity in 36% of tumours, the majority of which was rather focal and weak. Taken together, NKX3.1 expression showed a low sensitivity but a high specificity in our analysis. Methylome profiling on the other hand represents a sensitive, specific and reliable tool to support the diagnosis of MCS, particularly if only the round cell component is obtained in a biopsy and the diagnosis is not suspected. Furthermore, it can aid in confirming the diagnosis in case RNA sequencing for the HEY1::NCOA2 fusion transcript is not available.

CDX2 protein distribution in the gastric mucosa in chronic gastritis by a semi-quantitative index and its reproducibility

Introduction The Cdx2 gene provides an intestinal differentiation of epithelial cells and plays an oncosupressive role. An indirect method of the Cdx2 gene expression assessment is the immunohistochemical study of its product, the CDX2 protein. Therefore, the common approach to the immunohistochemical study of the CDX2 protein hasn’t been developed yet. A semi-quantitative CDX2 index based on the percentage of CDX2-positive cells in the tissue specimen, the staining intensity and an expression pattern has been proposed.The purpose of the study was to assess the reproducibility of the semi-quantitative CDX2 index calculation in chronic atrophic gastritis stages I-IV.Materials and methods 20 chronic atrophic gastritis cases (5 cases for each stage of the gastritis according to the Operative link for gastritis assessment system) were taken according to the Maastricht V protocol and examined by the immunohistochemical method (CDX2, clone EPR2764Y, ready to use). The reproducibility of the CDX2 semi-quantitative index was assessed by five pathologists. An agreement between observed raters was measured by the kappa statistics.Results The Cohen’s κ value is 0,8 for unweighted κ and 0,97 for weighted κ (extremely high level of agreement) for the semi-quantitative CDX2 index calculation.Discussion The least reproducible parameter used for the semi-quantitative CDX2 index calculation the percentage of CDX2-positive cells because of the subjective assessment. The Cohen’s weighted κ value was higher compared to the unweighted κ because of the close yet not similar CDX2 semi-quantitative index values calculated by pathologists.Conclusion The semi-quantitative CDX2 index can be used to rank CDX2 expression and has a high level of reproducibility.