Immunophenotypic classification regarding prognosis in peripheral T cell lymphoma, NOS, and nodal T follicular helper T cell lymphoma, angioimmunoblastic-type.

This study aimed to determine the clinicopathological predictive factors of peripheral T-cell lymphoma, not otherwise specified (PTCL, NOS), and nodal T-follicular helper cell lymphoma, angioimmunoblastic-type (nTFH, AI-type). In this single-centered, retrospective study, medical records of 59 patients who were diagnosed with PTCL, NOS, or nTFH, AI-type from March 2007 to September 2022 were reviewed. The clinicopathological variables, including immunohistochemistry(IHC) subgroups, distinguishing TBX21 from the GATA3 subgroups were analyzed. Overall, 28 patients (75.7%) in the TBX21 group were PTCL, NOS. There were 9 (24.3%) patients in the GATA3 group. In univariable analyses, lymphoma subtype, age, and performance status were associated with progression-free survival (PFS), and overall survival (OS). In multivariable analyses, lymphoma subtype, and performance status were related to PFS and OS (P = 0.012, P < 0.001, P = 0.006, and P < 0.001, respectively). The GATA3 subgroup tended to have a worse prognosis in univariable analyses; however, it became more insignificant in multivariable when lymphoma subtype and performance status were adjusted (P = 0.065, P = 0.180, P = 0.972, and P = 0.265, respectively). The double-positive group showed variable prognoses of better PFS and worse OS. PD-1 and PD-L1 were associated with the EBV in situ hybridization (P = 0.027, and P = 0.005), and PD-1 was associated with CD30 expression (P = 0.043). This study demonstrated the potential of IHC classification to predict prognosis for PTCL, NOS, as well as nTFH AI-type, although further validation is necessary. Treatments targeting CD30, PD-1, and PD-L1 appear promising for lymphoma treatment.

Characterization of Tbr2-expressing retinal ganglion cells.

The mammalian retina contains more than 40 retinal ganglion cell (RGC) subtypes based on their unique morphologies, functions, and molecular profiles. Among them, intrinsically photosensitive RGCs (ipRGCs) are the first specified RGC type emerging from a common retinal progenitor pool during development. Previous work has shown that T-box transcription factor T-brain 2 (Tbr2) is essential for the formation and maintenance of ipRGCs, and that Tbr2-expressing RGCs activate Opn4 expression upon native ipRGC ablation, suggesting that Tbr2+ RGCs contain a reservoir for ipRGCs. However, the identity of Tbr2+ RGCs has not been fully vetted. Here, using genetic sparse labeling and single cell recording, we showed that Tbr2-expressing retinal neurons include RGCs and a subset of GABAergic displaced amacrine cells (dACs). Most Tbr2+ RGCs are intrinsically photosensitive and morphologically resemble native ipRGCs with identical retinofugal projections. Tbr2+ RGCs also include a unique and rare Pou4f1-expressing OFF RGC subtype. Using a loss-of-function strategy, we have further demonstrated that Tbr2 is essential for the survival of these RGCs and dACs, as well as maintaining the expression of Opn4. These data set a strong foundation to study how Tbr2 regulates ipRGC development and survival, as well as the expression of molecular machinery regulating intrinsic photosensitivity.

The potential role of follicular helper T cells and helper T cells type 1 in Warthin tumour.

Warthin tumour (WT) is a benign tumour of the salivary gland that proliferates in both glandular epithelial and lymphoid tissue components, and rarely exhibits cystic changes. T follicular helper cells (Tfh) are involved in the formation and maintenance of germinal centres, the differentiation of B cells into plasma cells, and the maintenance of helper T cell type 2 (Th2)-dominant humoral immune responses. T-bet induces differentiation into helper T cell type 1 (Th1) by suppressing differentiation into Tfh and enhances cellular immune responses. The objective of this study was to enhance our understanding of the immune responses and relationship between Tfh and Th1 cells in patients with WTs. In this study, we classified WTs (n = 64) into solid-type (n = 25) and cyst-type (n = 39). We also performed immunostaining of the Tfh markers CXCR5 and CD40 L, and the Th1 marker T-bet for statistical analysis. The cyst-type exhibited significant atrophy of the germinal centre area (P = 0.0019), significantly fewer Tfh-positive lymphocytes in germinal centres (P < 0.0001), and significantly more T-bet-positive lymphocytes in the epithelium (P = 0.0017). We observed that Tfh were involved in the formation and maintenance of lymphoid follicles in WTs. In the cyst-type, Th2-dominant humoral immune responses were suppressed, and Th1-dominant cellular immune responses may have caused damage to tumour tissue.

Decreased infiltration of CD4+ Th1 cells indicates a good response to ursodeoxycholic acid (UDCA) in primary biliary cholangitis.

BACKGROUND: Primary biliary cholangitis (PBC) is characterized by nonsuppurative destructive cholangitis and is thought to be an autoimmune disorder. Currently, ursodeoxycholic acid (UDCA) is the only FDA approved first-line therapy for PBC, but up to nearly one-third of patients do not achieve a complete response to this treatment. Adaptive immune cells, including T cells and B cells, have been found in the portal tracts and the bile duct epithelium and play a role in the pathogenesis of PBC, but the importance of these cells for evaluating the therapeutic response to UDCA in PBC has not yet been studied. METHODS: In this study, we collected liver puncture biopsy specimens from 34 matched patients with PBC before and after UDCA treatment and investigated the relationship between the infiltration of adaptive immune cells and the treatment response to UDCA. The extent of immune cell infiltration was determined by immunohistochemical analysis. Responses were defined based on Paris-I criteria. RESULTS: After 1 year of treatment, 25/34 patients responded to UDCA treatment according to Paris-I criteria (responders), and 9/34 patients were nonresponders. Immunohistochemical analysis showed that UDCA responders exhibited significantly less CD4+ T cell infiltration after UDCA treatment than before (50.4 ± 7.5/HPF vs 30.0 ± 7.9/HPF, P = 0.002). In contrast, UDCA nonresponders exhibited significantly more CD4+ T cell infiltration after UDCA treatment than before (32.2 ± 8.0/HPF vs 75.0 ± 13.9/HPF, P = 0.045). Moreover, patients who exhibited a reduction in CD4+ T cell infiltration after UDCA treatment had a higher response rate than those that exhibited an increase in CD4+ T cell infiltration (85.7 % vs 53.8 %, P = 0.041). However, CD3+ T cell, CD8+ T cell, and CD20+ B cell infiltration was not significantly different before and after treatment in either UDCA responders or nonresponders. Furthermore, we found that the number of infiltrating T-bet+ Th1 cells was much lower after UDCA treatment than before in responders (10.5 ± 5.7/HPF vs. 5.16 ± 4.0/HPF, P = 0.0214) but much higher in nonresponders after treatment than before (1.89±1.2/HPF vs. 12.3±5.4/HPF, P = 0.043). However, there was no difference in the extent of GATA3+ Th2 or FOXP3+ Treg infiltration before and after treatment in either UDCA responders or nonresponders. CONCLUSION: Collectively, our results suggest that a decrease in the number of liver-infiltrating CD4+ Th1 cells is associated with a good response of PBC patients to UDCA treatment. Immunohistochemical analysis of CD4 and T-bet in PBC liver specimens may be a potential approach for evaluating the therapeutic response to UDCA.

Retinoic acid-responsive CD8 effector T cells are selectively increased in IL-23-rich tissue in gastrointestinal GVHD.

Gastrointestinal (GI) graft-versus-host disease (GVHD) is a major barrier in allogeneic hematopoietic stem cell transplantation (allo-HSCT). The metabolite retinoic acid (RA) potentiates GI-GVHD in mice via alloreactive T cells expressing the RA receptor-α (RARα), but the role of RA-responsive cells in human GI-GVHD remains undefined. Therefore, we used conventional and novel sequential immunostaining and flow cytometry to scrutinize RA-responsive T cells in tissues and blood of patients who had received allo-HSCT and to characterize the impact of RA on human T-cell alloresponses. Expression of RARα by human mononuclear cells was increased after exposure to RA. RARαhi mononuclear cells were increased in GI-GVHD tissue, contained more cellular RA-binding proteins, localized with tissue damage, and correlated with GVHD severity and mortality. By using a targeted candidate protein approach, we predicted the phenotype of RA-responsive T cells in the context of increased microenvironmental interleukin-23 (IL-23). Sequential immunostaining confirmed the presence of a population of RARαhi CD8 T cells with the predicted phenotype that coexpressed the effector T-cell transcription factor T-bet and the IL-23-specific receptor (IL-23R). These cells were increased in GI- but not skin-GVHD tissues and were also selectively expanded in the blood of patients with GI-GVHD. Finally, functional approaches demonstrated that RA predominantly increased alloreactive GI-tropic RARαhi CD8 effector T cells, including cells with the phenotype identified in vivo. IL-23-rich conditions potentiated this effect by selectively increasing ß7 integrin expression on CD8 effector T cells and reducing CD4 T cells with a regulatory cell phenotype. In summary, we have identified a population of RA-responsive effector T cells with a distinctive phenotype that is selectively expanded in human GI-GVHD and that represents a potential new therapeutic target.

Utility of GATA-3 Expression in the Analysis of Pituitary Neuroendocrine Tumour (PitNET) Transcription Factors.

With the introduction of the WHO 2017 classification of endocrine neoplasms, the use of the pituitary transcription factors PIT-1, Tpit and SF-1 has become the standard of care. However, immunohistochemistry for these transcription factors is not available in all institutions, and their reliability has been questioned. We read with interest the findings of Mete et al. that GATA-3 expression was detected in some pituitary neuroendocrine tumours (PitNET). We therefore sort to validate this in our large cohort of PitNETs. We searched the database of Royal North Shore Hospital for PitNETs between 1998 and 2012, constructed a tissue microarray and reclassified these entities based on their expression for PIT-1, Tpit and SF-1. We then scored the expression of GATA-3 immunohistochemistry on a scale of 0-2, where 0 was no staining, 1 was patchy or weak staining and 2 was strong and diffuse staining. 265 of 346 tumours were able to be classified into a specific tumour subtype, and 263 tumours had tissue available for GATA-3 immunohistochemistry. 89% of gonadotrophs and 93% of triple-negative tumours with expression for luteinising hormone and follicle-stimulating hormone were positive for GATA-3. In the triple-negative group, GATA-3 was positive in 1 mammosomatotroph and 80% of tumours with thyroid-stimulating hormone expression. In the triple-negative hormone-negative group, 21 of 33 tumours were positive (64%). The results demonstrate that GATA-3 is a useful marker to supplement the existing pituitary transcription factors, albeit slightly less sensitive and specific than previously reported. GATA-3 may be employed in addition to the current array of immunohistochemical transcription factors, especially in the resource poor setting. However, given its potential cross-reactivity with other entities of the Sella, positive staining should be interpreted with caution and in the morphological and clinical context.

Brachyury: Strategies for Drugging an Intractable Cancer Therapeutic Target.

New approaches to drug discovery are unlocking enormous therapeutic potential residing in cancer-specific molecules. Brachyury is emerging as an exciting new drug target for the rare bone cancer chordoma. Here, recent advances targeting Brachyury in chordoma are discussed and how these might open doors to the targeting of other, more common cancer types.

Clinical Parameters of Silent Corticotroph Adenomas With Positive and Negative Adrenocorticotropic Hormone Immunostaining: A Large Retrospective Single-Center Study of 105 Cases.

Purpose: To investigate the different clinical characteristics of silent corticotroph adenomas (SCAs) with positive and negative adrenocorticotropic hormone (ACTH) immunostaining, and to explore the value of pituitary-restricted transcription factor (Tpit) immunostaining for diagnosing SCAs. Methods: The clinical materials of patients with SCAs who had a typical pathological feature with positive Tpit immunostaining and positive/negative ACTH immunostaining, and without clinical features and biochemical evidence for Cushing's Syndrome in our center from April 2018 to March 2019 were analyzed retrospectively. The differences in clinical characteristics and surgical results between ACTH-positive and -negative SCAs were explored. Results: A total of one hundred and five patients (94.3% female) with SCAs were included. There were 66 SCAs with ACTH-negative (66/105, 62.9%), and 39 SCAs with ACTH-positive (39/105, 37.1%). Cases with ACTH-negative SCAs were more likely to have lower ACTH levels (27.5 ± 24.0 vs. 54.4 ± 58.6, P = 0.011), more multiple microcysts (81.8% vs. 61.5%, P = 0.022) and lower levels of Ki-67 expression (low expression rate 90.9% vs. 74.4%, P = 0.023). No statistical significant differences were observed between patients with ACTH-positive and -negative SCAs regarding gender (97.0% vs. 89.7%, P = 0.192), age (50.3 ± 10.3 vs. 49.0 ± 11.2, P = 0.543), surgical history (16.7% vs. 23.1%, P = 0.419), suprasellar extension (66.7% vs. 74.4%, P = 0.408), sphenoid sinus extension (51.5% vs. 56.4%, P = 0.627), cavernous sinus invasion (75.8% vs. 66.7%, P = 0.314), large cyst on Magnetic Resonance Imaging (MRI) (47.0% vs. 61.5%, P = 0.149), or gross total resection rate (42.4% vs. 51.3%, P = 0.379). Conclusions: ACTH-negative SCAs were observed to be more clinically silent and more likely to demonstrate multiple microcysts on MRI. The prevalence of SCAs, especially ACTH-negative SCAs, proved to be substantially underestimated and thus they should be given enough attention in consideration of the high aggressiveness of this subtype of refractory pituitary adenoma (PA).

DNA hypermethylation of Fgf16 and Tbx22 associated with cleft palate during palatal fusion.

OBJECTIVE: Cleft palate (CP) is a congenital birth defect caused by the failure of palatal fusion. Little is known about the potential role of DNA methylation in the pathogenesis of CP. This study aimed to explore the potential role of DNA methylation in the mechanism of CP. METHODOLOGY: We established an all-trans retinoic acid (ATRA)-induced CP model in C57BL/6J mice and used methylation-dependent restriction enzymes (MethylRAD, FspEI) combined with high-throughput sequencing (HiSeq X Ten) to compare genome-wide DNA methylation profiles of embryonic mouse palatal tissues, between embryos from ATRA-treated vs. untreated mice, at embryonic gestation day 14.5 (E14.5) (n=3 per group). To confirm differentially methylated levels of susceptible genes, real-time quantitative PCR (qPCR) was used to correlate expression of differentially methylated genes related to CP. RESULTS: We identified 196 differentially methylated genes, including 17,298 differentially methylated CCGG sites between ATRA-treated vs. untreated embryonic mouse palatal tissues (P<0.05, log2FC>1). The CP-related genes Fgf16 (P=0.008, log2FC=1.13) and Tbx22 (P=0.011, log2FC=1.64,) were hypermethylated. Analysis of Fgf16 and Tbx22, using Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG), identified 3 GO terms and 1 KEGG pathway functionally related to palatal fusion. The qPCR showed that changes in expression level negatively correlated with methylation levels. CONCLUSIONS: Taken together, these results suggest that hypermethylation of Fgf16 and Tbx22 is associated with decreased gene expression, which might be responsible for developmental failure of palatal fusion, eventually resulting in the formation of CP.

IFNγ induces epigenetic programming of human T-bethi B cells and promotes TLR7/8 and IL-21 induced differentiation.

Although B cells expressing the IFNγR or the IFNγ-inducible transcription factor T-bet promote autoimmunity in Systemic Lupus Erythematosus (SLE)-prone mouse models, the role for IFNγ signaling in human antibody responses is unknown. We show that elevated levels of IFNγ in SLE patients correlate with expansion of the T-bet expressing IgDnegCD27negCD11c+CXCR5neg (DN2) pre-antibody secreting cell (pre-ASC) subset. We demonstrate that naïve B cells form T-bethi pre-ASCs following stimulation with either Th1 cells or with IFNγ, IL-2, anti-Ig and TLR7/8 ligand and that IL-21 dependent ASC formation is significantly enhanced by IFNγ or IFNγ-producing T cells. IFNγ promotes ASC development by synergizing with IL-2 and TLR7/8 ligands to induce genome-wide epigenetic reprogramming of B cells, which results in increased chromatin accessibility surrounding IRF4 and BLIMP1 binding motifs and epigenetic remodeling of IL21R and PRDM1 loci. Finally, we show that IFNγ signals poise B cells to differentiate by increasing their responsiveness to IL-21.

Fine particulate matter (PM2.5) aggravates apoptosis of cigarette-inflamed bronchial epithelium in vivo and vitro.

Fine particulate matter (PM2.5) is an essential risk factor of chronic obstructive pulmonary disease (COPD). Recent studies showed weak association between PM2.5 and COPD incidence, but smokers who exposed to higher PM2.5 concentration had more opportunity to gain COPD. Cigarette smoking is the most important risk factor of COPD. Thus, we hypothesized: the role of PM2.5 played on cigarette-inflamed airways was more significant than normal airways. The study firstly established an animal model of C57BL/6J mice with cigarette smoke exposure and PM2.5 orotracheal administration. After calculating pathological scores, mean linear intercept and mean alveolar area, we found PM2.5 aggravated pathological injury of cigarette-inflamed lungs, but the injury on normal lungs was not significant. Meanwhile, inflammatory factors as T-bet, IFN-γ and IL-1α were tested using qRT-PCR and ELISA. The results showed PM2.5 aggravated inflammation of cigarette-inflamed lungs, but the effect on normal lungs was not significant. The most important pathogenesis of COPD is abnormal apoptosis in airway epithelium, due to oxidative stress following long-term exposure to cigarette smoke. Then, apoptotic responses were detected in lungs. TUNEL analysis demonstrated that PM2.5 promoted DNA fragmentation of cigarette-inflamed lungs, but the effect on normal lungs was not significant. Western-blot and immunohistochemistry showed caspase activated significantly in PM2.5-cigarette smoke exposed lungs and activated caspase 3 located mainly on bronchial epithelium. Next, human bronchial epithelial cells were cultured treated with cigarette smoke solution (CSS) with or without PM2.5. Z-VAD-FMK, a pan-caspase inhibitor, was used to suppress the activation of caspases. After analyzing cell viability, DNA fragmentation, mitochondrial activities and caspase activities, the results clarified that PM2.5 aggravated apoptosis in cigarette-inflamed bronchial epithelial cells and the responses could be suppressed by Z-VAD-FMK. Our results gave a new idea about the mechanism of PM2.5 on COPD and inferred cigarette-inflamed airways were more vulnerable to PM2.5 than normal airways.

DNA hypermethylation of Fgf16 and Tbx22 associated with cleft palate during palatal fusion

Abstract Objective: Cleft palate (CP) is a congenital birth defect caused by the failure of palatal fusion. Little is known about the potential role of DNA methylation in the pathogenesis of CP. This study aimed to explore the potential role of DNA methylation in the mechanism of CP. Methodology: We established an all-trans retinoic acid (ATRA)-induced CP model in C57BL/6J mice and used methylation-dependent restriction enzymes (MethylRAD, FspEI) combined with high-throughput sequencing (HiSeq X Ten) to compare genome-wide DNA methylation profiles of embryonic mouse palatal tissues, between embryos from ATRA-treated vs. untreated mice, at embryonic gestation day 14.5 (E14.5) (n=3 per group). To confirm differentially methylated levels of susceptible genes, real-time quantitative PCR (qPCR) was used to correlate expression of differentially methylated genes related to CP. Results: We identified 196 differentially methylated genes, including 17,298 differentially methylated CCGG sites between ATRA-treated vs. untreated embryonic mouse palatal tissues (P<0.05, log2FC>1). The CP-related genes Fgf16 (P=0.008, log2FC=1.13) and Tbx22 (P=0.011, log2FC=1.64,) were hypermethylated. Analysis of Fgf16 and Tbx22, using Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG), identified 3 GO terms and 1 KEGG pathway functionally related to palatal fusion. The qPCR showed that changes in expression level negatively correlated with methylation levels. Conclusions: Taken together, these results suggest that hypermethylation of Fgf16 and Tbx22 is associated with decreased gene expression, which might be responsible for developmental failure of palatal fusion, eventually resulting in the formation of CP.

Tumoral BRD4 expression in lymph node-negative breast cancer: association with T-bet+ tumor-infiltrating lymphocytes and disease-free survival.

BACKGROUND: We previously observed that T-bet+ tumor-infiltrating T lymphocytes (T-bet+ TILs) in primary breast tumors were associated with adverse clinicopathological features, yet favorable clinical outcome. We identified BRD4 (Bromodomain-Containing Protein 4), a member of the  Bromodomain and Extra Terminal domain (BET) family, as a gene that distinguished T-bet+/high and T-bet-/low tumors. In clinical studies, BET inhibitors have been shown to suppress inflammation in various cancers, suggesting a potential link between BRD4 and immune infiltration in cancer. Hence, we examined the BRD4 expression and clinicopathological features of breast cancer. METHODS: The cohort consisted of a prospectively ascertained consecutive series of women with axillary node-negative breast cancer with long follow-up. Gene expression microarray data were used to detect mRNAs differentially expressed between T-bet+/high (n = 6) and T-bet-/low (n = 41) tumors. Tissue microarrays (TMAs) constructed from tumors of 612 women were used to quantify expression of BRD4 by immunohistochemistry, which was analyzed for its association with T-bet+ TILs, Jagged1, clinicopathological features, and disease-free survival. RESULTS: Microarray analysis indicated that BRD4 mRNA expression was up to 44-fold higher in T-bet+/high tumors compared to T-bet-/low tumors (p = 5.38E-05). Immunohistochemical expression of BRD4 in cancer cells was also shown to be associated with T-bet+ TILs (p = 0.0415) as well as with Jagged1 mRNA and protein expression (p = 0.0171, 0.0010 respectively). BRD4 expression correlated with larger tumor size (p = 0.0049), pre-menopausal status (p = 0.0018), and high Ki-67 proliferative index (p = 0.0009). Women with high tumoral BRD4 expression in the absence of T-bet+ TILs exhibited a significantly poorer outcome (log rank test p = 0.0165) relative to other subgroups. CONCLUSIONS: The association of BRD4 expression with T-bet+ TILs, and T-bet+ TIL-dependent disease-free survival suggests a potential link between BRD4-mediated tumor development and tumor immune surveillance, possibly through BRD4's regulation of Jagged1 signaling pathways. Further understanding BRD4's role in different immune contexts may help to identify an appropriate subset of breast cancer patients who may benefit from BET inhibitors without the risk of diminishing the anti-tumoral immune activity.

Clinicopathological and immunohistochemical analysis of plasma cell-rich rejection in renal transplantation: Involvement of intratubular Th1/Th2 balance in plasma cell enrichment.

AIM: Plasma cell-rich rejection (PCRR) has been considered a subtype of acute T-cell-mediated rejection (ATCR). However, PCRR is recognized as refractory rejection and different from ATCR in various ways. In order to elucidate the pathogenesis of PCRR, we analysed PCRR clinicopathologically and immunohistochemically by comparing it with ATCR. METHODS: Twelve cases of PCRR (PCRRs) and 22 cases of usual ATCR (ATCRs) diagnosed at our hospital between January 2008 and March 2017 were included. Between PCRRs and ATCRs, we compared clinical data, Banff classification, graft outcome and the total sum number of T-bet- and GATA3-positive lymphocytes infiltrating in tubular epithelium using immunohistochemistry. RESULTS: Plasma cell-rich rejections occurred later than ATCRs (median time after transplantation 1340.5 days vs. 52.5 days). Serum creatinine levels at discharge after treatment were significantly higher in PCRRs than in ATCRs (median 2.38 vs. 1.65 mg/dL). Cumulative rate of graft loss was significantly higher in PCRRs than in ATCRs (1-, 2- and 5-year: 26.7%, 51.1% and 51.1% vs. 0%, 0% and 17.5%). For profiles of Th1 and Th2, we found significantly lower ratio of T-bet/GATA3-positive lymphocytes in PCRRs compared with ATCRs. CONCLUSION: This study suggests that PCRR is more refractory than ATCR and there are significant differences in populations of helper T-cell subsets between them. We consider helper T-cell subset analysis valuable for developing new treatment strategies for PCRR.

Chordoma: Immunohistochemical Analysis of Brachury.

AIM: Chordomas are rare, slow growing but locally aggressive malignancies of the axial skeleton. Skull base chordomas, due to their intricate anatomical localization, pose significant challenges to managing physicians. In classical and chondroid chordomas, the disease course cannot be reliably determined using only morphological criteria. Brachyury (T Gene) was shown to play a central role in chordoma pathogenesis and several studies also showed that this gene also carries potential as a prognostic biomarker. This study aims to correlate Brachyury expression with the clinical course in surgically treated skull base chordomas. MATERIAL AND METHODS: Chordoma tumor samples from 14 patients with skull base chordomas, diagnosed using histopathological and immunohistochemistry criteria (epithelial membrane antigen (EMA), S100, pan cytokeratin (panCK)) were retrospectively analyzed for Brachyury expression using immunohistochemistry. Brachyury expression was graded using a 4 point semi-quantitative scoring system. Focal (grade II) and diffuse staining (grade III) were considered as overexpression. Patient recurrence-free survival and total survival were compared between Brachyury overexpressing and non-overexpressing groups using Kaplan-Meier survival analysis. RESULTS: Among the stained tumor samples, 85.7% were positive for brachyury expression. In both groups, there was one sample that was negative. We did not observe any significant difference among the groups for staining, grade and percentage of brachyury positive cells. CONCLUSION: Brachyury expression in tumor samples is not a sensitive indicator of prognosis in chordomas.

Parosteal extra-axial chordoma of the second metacarpal bone: a case report with literature review.

Extra-axial chordoma is a chordoma that occurs in non-axial locations. It is a very rare tumor, with 20 cases reported to date; 14 in bone and six in soft tissue. Of the 14 skeletal extra-axial chordomas, ten were intramedullary and four were intracortical. We report the first case of parosteal extra-axial chordoma arising in the second metacarpal bone, expressing brachyury on immunohistochemical analysis, and describe the pathologic and radiologic findings. We suggest that extra-axial chordoma can occur in parosteal bone lesions or the hand, without features of bone distribution or bone-specific sites.

TBX19 is overexpressed in colorectal cancer and associated with lymph node metastasis.

The T-box 19 (TBX19) gene encodes a transcription factor characterized by a highly conserved DNA-binding motif (T-box). Recent studies have revealed that TBX19 has been identified as one of the genes activated by KRAS mutations, and is upregulated in colon adenoma. These results indicate that TBX19 may work as an oncogene in colorectal cancer (CRC). However, the expression and role of TBX19 have yet to be investigated. Here, we investigated TBX19 mRNA and protein expressions in colon cancer cells or surgically resected CRC. We found that TBX19 mRNA expression was significantly increased in tumorous tissues compared to that in non-tumorous tissues, and increased TBX19 mRNA expression was associated with positive lymph node metastasis in our cohort. The expression of TBX19 mRNA was not correlated with that of TBX19 protein in tissue sample taken from the CRC patients. Moreover, TBX19 showed positive staining even in the normal colonic tissues and the adjacent non-tumorous tissues. These results suggest that the expression of TBX19 protein is not correlated with the expression of TBX19 mRNA. In addition, our results promote further investigations into the impact of TBX19 upregulation on colorectal carcinogenesis, as well as the underlying mechanisms.

Clinicopathologic features of four rare types of chordomas, confirmed by brachyury immunostaining.

BACKGROUND: A wide clinicopathologic spectrum of a chordoma exists. Brachyury constitutes as its most useful diagnostic immunohistochemical (IHC) marker. METHODS: During a 7-year-period, 4 unusual histopathologic types of chordomas were identified. Immunohistochemistry was performed by polymer technique. RESULTS: Clinicopathologic features of the 4 cases are as follows: Cases 1 and 2: Two tumors occurred in the sacrococcygeal and lumbosacral regions of a 42-year-old male and a 34-year-old female, respectively. Histopathologic examination showed areas of classical chordoma; juxtaposed to a high-grade, spindle cell sarcoma. By IHC, cytokeratin (CK), epithelial membrane antigen (EMA), S-100 protein, and brachyury were found to be distinctly positive in the differentiated chordomatous areas. Both these cases were diagnosed as dedifferentiated chordomas. The first patient, postresection and adjuvant radiation therapy (RT), died after 14 months of therapy. Case 3: A 58-year-old male presented with pain in his sacral region and urinary incontinence. Imaging disclosed a sacral mass. Histopathologic examination showed physaliphorous cells intimately admixed with, markedly pleomorphic cells, scattered mitotic figures, and focal tumor necrosis. By IHC, the tumor cells were positive for CK, AE1/AE3, S-100 protein, brachyury, and INI1/SMARCB1. The diagnosis of a poorly differentiated chordoma was offered. Despite surgical resection and adjuvant RT, the patient died within 18 months. Case 4: A 58-year-old male presented with a soft tissue lesion in his left leg. Histopathologic examination showed physaliphorous cells, embedded in a myxohyaline stroma. By IHC, the tumor cells were positive for EMA, S-100 protein, brachyury, and INI1. Diagnosis of an extra-axial, soft tissue chordoma was offered. CONCLUSIONS: These four unusual chordomas, confirmed by brachyury immunoexpression, constitute as one of the first such documentation from our country, revealing a wide clinicopathologic spectrum of chordomas. Dedifferentiated and poorly differentiated chordomas are associated with an aggressive clinical course. Further diagnostic implications are discussed herewith.