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Genome-wide association studies and experimental mouse models implicate the MIB1 and GATA6 genes in congenital heart disease (CHD). Their close physical proximity and conserved synteny suggest that these two genes might be involved in analogous cardiac developmental processes. Heterozygous Gata6 loss-of-function mutations alone or humanized Mib1 mutations in a NOTCH1-sensitized genetic background cause bicuspid aortic valve (BAV) and a membranous ventricular septal defect (VSD), consistent with MIB1 and NOTCH1 functioning in the same pathway. To determine if MIB1-NOTCH and GATA6 interact in valvular and septal development, we generated compound heterozygote mice carrying different Mib1 missense (Mib1K735R and Mib1V943F) or nonsense (Mib1R530X) mutations with the Gata6STOP/+ heterozygous null mutation. Combining Mib1R530X/+ or Mib1K735R/+ with Gata6STOP/+ does not affect Gata6STOP/+ single mutant phenotypes. In contrast, combining Mib1V943F/+ with Gata6STOP/+ decreases the incidence of BAV and VSD by 50%, suggesting a suppressive effect of Mib1V943F/+ on Gata6STOP/+. Transcriptomic and functional analyses revealed that while the EMT pathway term is depleted in the Gata6STOP/+ mutant, introducing the Mib1V943F variant robustly enriches this term, consistent with the Mib1V943F/+ phenotypic suppression of Gata6STOP/+. Interestingly, combined Notch1 and Gata6 insufficiency led to a nearly fully penetrant VSD but did not affect the BAV phenotype, underscoring the complex functional relationship between MIB1, NOTCH, and GATA6 in valvular and septal development.

Sanggenon C (SC), a herbal flavonoid extracted from Cortex Mori, has been mentioned to possess more than one treasured organic properties. However, the molecular mechanism of its anti‐tumor impact in glioblastoma (GBM) remains unclear. In this study, we reported that SC displayed a GBM‐suppressing impact in vitro and in vivo with no apparent organ toxicity. SC dramatically suppressed cell proliferation‐induced cell apoptosis in GBM cells. Mechanistically, we unveiled that SC modulated the protein expression of death associated protain kinase 1 (DAPK1) by controlling the ubiquitination and degradation of DAPK1. Quantitative proteomic and Western blot analyses showed that SC improved DAPK1 protein degradation via decreasing the expression of E3 ubiquitin ligase Mindbomb 1 (MIB1). More importantly, the effects of SC on cell proliferation and apoptosis of GBM cells have been in part reversed through DAPK1 downregulation or MIB1 overexpression, respectively. These results indicated that SC might suppress cell proliferation and induce cell apoptosis by decreasing MIB1‐mediated DAPK1 degradation. Furthermore, we found that SC acted synergistically with temozolomide (TMZ), an anti‐cancer drug used in GBM, resulting in elevated chemotherapeutic sensitivity of GBM to TMZ. Collectively, our data suggest that SC might be a promising anti‐cancer agent for GBM therapy. [ABSTRACT FROM AUTHOR]

Breast cancer is one of the most common invasive cancers among women. The leading cause of difficulty in treating breast cancer patients is metastasis. Because cell migration is closely related to breast cancer metastasis, elucidating the detailed mechanism by which breast cancer cells promote their migration is crucial for improving the prognosis of patients. In this study, we investigated the relationship between breast cancer cell migration and Mind bomb1 (MIB1), an E3 ubiquitin ligase. We found that the downregulation of MIB1 promotes the cell migration of MCF7, a breast cancer-derived cell line. Furthermore, knockdown of MIB1 caused a reduction in CTNND1 and thereby impaired E-cadherin membrane localization in the cell boundary region. Taken together, our data suggest that MIB1 might play a role in suppressing breast cancer cell migration. • MIB1 expression is correlated with the worse prognosis of breast cancer patients. • MIB1 suppresses cell migration in breast cancer cell lines. • MIB1 regulates CTNND1 protein level and E-cadherin localization in the cell boundary. [ABSTRACT FROM AUTHOR]

Background: Acute myeloid leukemia (AML) is a complex disease characterized by clonal expansion of undifferentiated myeloid precursors, resulting in impaired hematopoiesis and bone marrow failure. Different genetic and environmental factors are believed to be involved in the pathogenesis of AML. Notch signaling with a tumor suppressing plays a role in myeloproliferative disorder and is a negative regulator of myeloid progenitor commitment. Crosstalk between Notch signaling and CXCR4 axis is a matter of debate in AML.Method: In the current case-control study, we evaluated the expression level of CXCR4, JAG1, and MIB1, which are all involved or related to Notch signaling in adult AML patients. Blood samples were obtained from 25 AML and 17 healthy individuals and the expression level of the selected genes was evaluated via the real-time polymerase chain reaction.Results: Our results revealed the increased expression of JAG1, but decreased expression of CXCR4 in AML patients in Iranian population of AML patients. Moreover, some gender-associated effects on the expression of JAG1 and CXCR4 were detected, which may be related to sex hormones. The expression level of MIB1 did not change significantly. The correlation analysis showed no correlations between the age of the patients and the expression levels of the genes.Conclusion: Herein, for the first time, we suggested some new evidence regarding the complex role of Notch signaling-related genes (CXCR4 and JAG1) in the pathogenesis of AML in Iranian patients.

Abstract Background Acute pancreatitis is the sudden inflammation of the pancreas. Severe cases of acute pancreatitis are potentially fatal and have no specific treatment available. Premature trypsinogen activation could initiate acute pancreatitis. However, the mechanism underlying premature trypsinogen activation is not fully understood. Methods In this research, a primary pancreatic acinar cell or mouse acute pancreatitis model was constructed. The effect of acid ceramidase (ASAH1), which is responsible for sphingosine production, was investigated in trypsinogen activation in vitro and in vivo. Meanwhile, the proteins regulating ASAH1 or binding to sphingosine were also detected by co-immunoprecipitation followed by mass spectrometry. Results The results showed that ASAH1 increased in acute pancreatitis. Increased ASAH1 promoted the activation of trypsinogen and cathepsin B. On the contrary, ASAH1 downregulation inhibited trypsinogen and cathepsin B. Meanwhile, ASAH1 regulated the activity of trypsin and cathepsin B through sphingosine. Additionally, E3 ligase Mind bomb homolog 1 (MIB1) decreased in acute pancreatitis resulting in the decreased binding between MIB1 and ASAH1. Exogenous MIB1 diminished the elevation in trypsin activity induced by acute pancreatitis inducer. ASAH1 increased owing to the inhibition of the proteasome degradation by MIB1. In acute pancreatitis, sphingosine was found to bind to pyruvate kinase. Pyruvate kinase activation could reduce trypsinogen activation and mitochondrial reactive oxygen species (ROS) production induced by sphingosine. Conclusions In conclusion, during the process of acute pancreatitis, MIB1 downregulation led to ASAH1 upregulation, resulting in pyruvate kinase inhibition, followed by trypsinogen activation.

Abstract Sanggenon C (SC), a herbal flavonoid extracted from Cortex Mori, has been mentioned to possess more than one treasured organic properties. However, the molecular mechanism of its anti‐tumor impact in glioblastoma (GBM) remains unclear. In this study, we reported that SC displayed a GBM‐suppressing impact in vitro and in vivo with no apparent organ toxicity. SC dramatically suppressed cell proliferation‐induced cell apoptosis in GBM cells. Mechanistically, we unveiled that SC modulated the protein expression of death associated protain kinase 1 (DAPK1) by controlling the ubiquitination and degradation of DAPK1. Quantitative proteomic and Western blot analyses showed that SC improved DAPK1 protein degradation via decreasing the expression of E3 ubiquitin ligase Mindbomb 1 (MIB1). More importantly, the effects of SC on cell proliferation and apoptosis of GBM cells have been in part reversed through DAPK1 downregulation or MIB1 overexpression, respectively. These results indicated that SC might suppress cell proliferation and induce cell apoptosis by decreasing MIB1‐mediated DAPK1 degradation. Furthermore, we found that SC acted synergistically with temozolomide (TMZ), an anti‐cancer drug used in GBM, resulting in elevated chemotherapeutic sensitivity of GBM to TMZ. Collectively, our data suggest that SC might be a promising anti‐cancer agent for GBM therapy.

Thoracic aortic aneurysms (TAA) consist of abnormal dilation or the widening of a portion of the ascending aorta, due to weakness or destructuring of the walls of the vessel and are potentially lethal. The congenital bicuspid aortic valve (BAV) is considered a risk factor for the development of TAA because asymmetric blood flow through the bicuspid aortic valve detrimentally influences the wall of the ascending aorta. NOTCH1 mutations have been associated with non-syndromic TAAs as a consequence of BAV, but little is known regarding its haploinsufficiency and its relationship with connective tissue abnormalities. We report two cases in which there is clear evidence that alterations in the NOTCH1 gene are the cause of TAA in the absence of BAV. On the one hand, we describe a 117 Kb deletion that includes a large part of the NOTCH1 gene and no other coding genes, suggesting that haploinsufficiency can be considered a pathogenic mechanism for this gene associated with TAA. In addition, we describe two brothers who carry two variants, one in the NOTCH1 gene and another in the MIB1 gene, corroborating the involvement of different genes of the Notch pathway in aortic pathology. [ABSTRACT FROM AUTHOR]

Abstract Background Circular RNAs (circRNAs) regulate various biological activities and have been shown to play crucial roles in hepatocellular carcinoma (HCC) progression. However, only a few coding circRNAs have been identified in cancers, and their roles in HCC remain elusive. This study aimed to identify coding circRNAs and explore their function in HCC. Methods CircMAP3K4 was selected from the CIRCpedia database. We performed a series of experiments to determine the characteristics and coding capacity of circMAP3K4. We then used in vivo and in vitro assays to investigate the biological function and mechanism of circMAP3K4 and its protein product, circMAP3K4-455aa, in HCC. Results We found circMAP3K4 to be an upregulated circRNA with coding potential in HCC. IGF2BP1 recognized the circMAP3K4 N6-methyladenosine modification and promoted its translation into circMAP3K4-455aa. Functionally, circMAP3K4-455aa prevented cisplatin-induced apoptosis in HCC cells by interacting with AIF, thus protecting AIF from cleavage and decreasing its nuclear distribution. Moreover, circMAP3K4-455aa was degraded through the ubiquitin–proteasome E3 ligase MIB1 pathway. Clinically, a high level of circMAP3K4 is an independent prognostic factor for adverse overall survival and adverse disease-free survival of HCC patients. Conclusions CircMAP3K4 is a highly expressed circRNA in HCC. Driven by m6A modification, circMAP3K4 encoded circMAP3K4-455aa, protected HCC cells from cisplatin exposure, and predicted worse prognosis of HCC patients. Targeting circMAP3K4-455aa may provide a new therapeutic strategy for HCC patients, especially for those with chemoresistance. Graphical Abstract CircMAP3K4 is a highly expressed circRNA in HCC. Driven by m6A modification, IGF2BP1 facilitates circMAP3K4 peptide translation, then the circMAP3K4 peptide inhibits AIF cleavage and nuclear distribution, preventing HCC cells from cell death under stress and promoting HCC progression.

Background: The complex genetics underlying human cardiac disease is evidenced by its heterogenous manifestation, multigenic basis, and sporadic occurrence. These features have hampered disease modeling and mechanistic understanding. Here, we show that 2 structural cardiac diseases, left ventricular noncompaction (LVNC) and bicuspid aortic valve, can be caused by a set of inherited heterozygous gene mutations affecting the NOTCH ligand regulator MIB1 (MINDBOMB1) and cosegregating genes.Methods: We used CRISPR-Cas9 gene editing to generate mice harboring a nonsense or a missense MIB1 mutation that are both found in LVNC families. We also generated mice separately carrying these MIB1 mutations plus 5 additional cosegregating variants in the ASXL3, APCDD1, TMX3, CEP192, and BCL7A genes identified in these LVNC families by whole exome sequencing. Histological, developmental, and functional analyses of these mouse models were carried out by echocardiography and cardiac magnetic resonance imaging, together with gene expression profiling by RNA sequencing of both selected engineered mouse models and human induced pluripotent stem cell-derived cardiomyocytes. Potential biochemical interactions were assayed in vitro by coimmunoprecipitation and Western blot.Results: Mice homozygous for the MIB1 nonsense mutation did not survive, and the mutation caused LVNC only in heteroallelic combination with a conditional allele inactivated in the myocardium. The heterozygous MIB1 missense allele leads to bicuspid aortic valve in a NOTCH-sensitized genetic background. These data suggest that development of LVNC is influenced by genetic modifiers present in affected families, whereas valve defects are highly sensitive to NOTCH haploinsufficiency. Whole exome sequencing of LVNC families revealed single-nucleotide gene variants of ASXL3, APCDD1, TMX3, CEP192, and BCL7A cosegregating with the MIB1 mutations and LVNC. In experiments with mice harboring the orthologous variants on the corresponding Mib1 backgrounds, triple heterozygous Mib1 Apcdd1 Asxl3 mice showed LVNC, whereas quadruple heterozygous Mib1 Cep192 Tmx3;Bcl7a mice developed bicuspid aortic valve and other valve-associated defects. Biochemical analysis suggested interactions between CEP192, BCL7A, and NOTCH. Gene expression profiling of mutant mouse hearts and human induced pluripotent stem cell-derived cardiomyocytes revealed increased cardiomyocyte proliferation and defective morphological and metabolic maturation.Conclusions: These findings reveal a shared genetic substrate underlying LVNC and bicuspid aortic valve in which MIB1-NOTCH variants plays a crucial role in heterozygous combination with cosegregating genetic modifiers. [ABSTRACT FROM AUTHOR]

It has been shown that multidrug and toxic compound extrusion/detoxification (MATE/DTX) family transporters are involved in the regulation of plant development and stress response. Here, we characterized the mini body1 (mib1) mutants in mungbean, which gave rise to increased branches, pentafoliate compound leaves, and shortened pods. Map-based cloning revealed that MIB1 encoded a MATE/DTX family protein in mungbean. qRTPCR analysis showed that MIB1 was expressed in all tissues of mungbean, with the highest expression level in the young inflorescence. Complementation assays in Escherichia coli revealed that MIB1 potentially acted as a MATE/DTX transporter in mungbean. It was found that overexpression of the MIB1 gene partially rescued the shortened pod phenotype of the Arabidopsis dtx54 mutant. Transcriptomic analysis of the shoot buds and young pods revealed that the expression levels of several genes involved in the phytohormone pathway and developmental regulators were altered in the mib1 mutants. Our results suggested that MIB1 plays a key role in the control of plant architecture establishment in mungbean. [ABSTRACT FROM AUTHOR]

As one of the few animals with variation in the number of rib pairs (RIB), the pig is a good model to study the mechanism of RIB regulation. Quantitative trait loci (QTL) for porcine RIB are present on Sus scrofa chromosome 7 (SSC7). Although several candidate genes exist in this QTL region on SSC7, the causal gene has yet to be verified. Beijing Black pig with 14–17 RIB is a good population for candidate gene mining and 1104 individuals were genotyped using the Illumina Porcine 50K BeadChip. A total of 14 SNPs from 95.49 to 97.78 Mb on SSC7 showed genome‐wide significant association with RIB. On SSC7, a locuszoom plot using pairwise linkage disequilibrium displayed the narrowest linkage region encompassing only two genes, ABCD4 and VRTN. In mice, a transcriptome expression profile was obtained using three embryos at E9.5 (the critical period for rib formation). ABCD4 was highly expressed, but no expression of VRTN was detected. On SSC6, there were four genome‐wide significant SNPs from 106.42 to 106.92 Mb associated with RIB. GREB1L and MIB1, in this region, were regarded as novel candidate genes. These results revealed a crucial candidate causal gene on SSC7 and novel genes on SSC6 for rib number and provided interesting new insights into its genetic basis. [ABSTRACT FROM AUTHOR]

Background: Acute myeloid leukemia (AML) is a complex disease characterized by clonal expansion of undifferentiated myeloid precursors, resulting in impaired hematopoiesis and bone marrow failure. Different genetic and environmental factors are believed to be involved in the pathogenesis of AML. Notch signaling with a tumor suppressing plays a role in myeloproliferative disorder and is a negative regulator of myeloid progenitor commitment. Crosstalk between Notch signaling and CXCR4 axis is a matter of debate in AML. Method: In the current case-control study, we evaluated the expression level of CXCR4, JAG1, and MIB1, which are all involved or related to Notch signaling in adult AML patients. Blood samples were obtained from 25 AML and 17 healthy individuals and the expression level of the selected genes was evaluated via the real-time polymerase chain reaction. Results: Our results revealed the increased expression of JAG1, but decreased expression of CXCR4 in AML patients in Iranian population of AML patients. Moreover, some gender-associated effects on the expression of JAG1 and CXCR4 were detected, which may be related to sex hormones. The expression level of MIB1 did not change significantly. The correlation analysis showed no correlations between the age of the patients and the expression levels of the genes. Conclusion: Herein, for the first time, we suggested some new evidence regarding the complex role of Notch signaling-related genes (CXCR4 and JAG1) in the pathogenesis of AML in Iranian patients. [ABSTRACT FROM AUTHOR]

Despite recent progress in cancer treatment, the prognosis of patients with pancreatic cancer still remains poor. Pancreatic tumors are reported to display high molecular heterogeneity. Elucidating the molecular mechanisms underlying pancreatic cancer progression is essential for improving patient treatment and survival. The overexpression of E3 ubiquitin ligase mind bomb 1 (MIB1) was previously described in pancreatic cancer cells, where it enhanced tumor cell proliferation. However, the role of MIB1 in pancreatic cancer progression remains elusive. In the present study, we confirmed that MIB1 expression is elevated in pancreatic cancer tissues and that high levels of MIB associate with unfavorable prognosis. Overexpression of MIB1 enhanced proliferation and invasion of pancreatic cancer cells both in vitro and in vivo. We further investigated the molecular mechanisms downstream of MIB1 and observed for the first time that MIB1 targets suppressor of tumorigenicity 7 protein (ST7), previously described as suppressor of tumorigenicity, for proteasomal degradation. Furthermore, we found that ST7 suppressed tumor growth by downregulating IQ motif containing GTPase activating protein 1 (IQGAP1) in pancreatic tumor cells. Thus, these data show that MIB1 promotes pancreatic cancer progression by inducing ST7 degradation followed by downregulation of IQGAP1 in pancreatic cancer cells. In conclusion, our research shows that the MIB1/ST7/IQGAP1 axis is essential for pancreatic cancer progression, and MIB1 inhibition may serve as a novel therapeutic strategy in patients with pancreatic cancer.

Abstract Background The dorsal domain of the neural tube is an excellent model to investigate the generation of complexity during embryonic development. It is a highly dynamic and multifaceted region being first transiently populated by prospective neural crest (NC) cells that sequentially emigrate to generate most of the peripheral nervous system. Subsequently, it becomes the definitive roof plate (RP) of the central nervous system. The RP, in turn, constitutes a patterning center for dorsal interneuron development. The factors underlying establishment of the definitive RP and its segregation from NC and dorsal interneurons are currently unknown. Results We performed a transcriptome analysis at trunk levels of quail embryos comparing the dorsal neural tube at premigratory NC and RP stages. This unraveled molecular heterogeneity between NC and RP stages, and within the RP itself. By implementing these genes, we asked whether Notch signaling is involved in RP development. First, we observed that Notch is active at the RP-interneuron interface. Furthermore, gain and loss of Notch function in quail and mouse embryos, respectively, revealed no effect on early NC behavior. Constitutive Notch activation caused a local downregulation of RP markers with a concomitant development of dI1 interneurons, as well as an ectopic upregulation of RP markers in the interneuron domain. Reciprocally, in mice lacking Notch activity, both the RP and dI1 interneurons failed to form and this was associated with expansion of the dI2 population. Conclusions Collectively, our results offer a new resource for defining specific cell types, and provide evidence that Notch is required to establish the definitive RP, and to determine the choice between RP and interneuron fates, but not the segregation of RP from NC.

Abstract Objective Vestibular schwannomas (VS) are benign slow growing tumors arising from the vestibular nerve. The role of cyclooxygenase 2 (COX2) in tumor development of growth has been addressed in a few studies with contradictory results and suggestions. We recently analyzed the immunohistochemical expression of COX2 in 1044 VS samples and described an association of higher COX2 expression with proliferation but found no influence by regular intake of acetylsalicylic acid. We now collected volumetric radiographic data of the preoperative tumor volume and growth to further test the role of COX2 in VS growth. Methods Preoperative images of 898 primary sporadic vestibular schwannomas were assessed, and sufficient preoperative imaging was used for the volumetric measurement preoperative tumor volume (n = 747) and preoperative relative tumor growth (n = 171). Clinical parameters and results of the immunohistochemical expression of COX2 and MIB1 in resected tumor tissue samples were obtained from our prior study. ANOVA, CART-analysis and multivariate nominal logistic regression were used for statistical analysis. Results Larger preoperative tumor volumes were observed with tumors of younger patients (p = 0.0288) and with higher COX2 expression scores (p

Background: Circular RNAs (circRNAs) regulate various biological activities and have been shown to play crucial roles in hepatocellular carcinoma (HCC) progression. However, only a few coding circRNAs have been identified in cancers, and their roles in HCC remain elusive. This study aimed to identify coding circRNAs and explore their function in HCC. Methods: CircMAP3K4 was selected from the CIRCpedia database. We performed a series of experiments to determine the characteristics and coding capacity of circMAP3K4. We then used in vivo and in vitro assays to investigate the biological function and mechanism of circMAP3K4 and its protein product, circMAP3K4-455aa, in HCC. Results: We found circMAP3K4 to be an upregulated circRNA with coding potential in HCC. IGF2BP1 recognized the circMAP3K4 N6-methyladenosine modification and promoted its translation into circMAP3K4-455aa. Functionally, circMAP3K4-455aa prevented cisplatin-induced apoptosis in HCC cells by interacting with AIF, thus protecting AIF from cleavage and decreasing its nuclear distribution. Moreover, circMAP3K4-455aa was degraded through the ubiquitin–proteasome E3 ligase MIB1 pathway. Clinically, a high level of circMAP3K4 is an independent prognostic factor for adverse overall survival and adverse disease-free survival of HCC patients. Conclusions: CircMAP3K4 is a highly expressed circRNA in HCC. Driven by m6A modification, circMAP3K4 encoded circMAP3K4-455aa, protected HCC cells from cisplatin exposure, and predicted worse prognosis of HCC patients. Targeting circMAP3K4-455aa may provide a new therapeutic strategy for HCC patients, especially for those with chemoresistance. CircMAP3K4 is a highly expressed circRNA in HCC. Driven by m6A modification, IGF2BP1 facilitates circMAP3K4 peptide translation, then the circMAP3K4 peptide inhibits AIF cleavage and nuclear distribution, preventing HCC cells from cell death under stress and promoting HCC progression. [ABSTRACT FROM AUTHOR]

Parkinson's disease remains one of the leading neurodegenerative diseases in developed countries. Despite well-defined symptomology and pathology, the complexity of Parkinson's disease prevents a full understanding of its etiological mechanism. Mechanistically, α-synuclein misfolding and aggregation appear to be central for disease progression, but mitochondrial dysfunction, dysfunctional protein clearance and ubiquitin/proteasome systems, and neuroinflammation have also been associated with Parkinson's disease. Particularly, neuroinflammation, which was initially thought to be a side effect of Parkinson's disease pathogenesis, has now been recognized as driver of Parkinson's disease exacerbation. Next-generation sequencing has been used to identify a plethora of long noncoding RNAs (lncRNA) with important transcriptional regulatory functions. Moreover, a myriad of lncRNAs are known to be regulators of inflammatory signaling and neurodegenerative diseases, including IL-1β secretion and Parkinson's disease. Here, LncZFAS1 was identified as a regulator of inflammasome activation, and pyroptosis in human neuroblast SH-SY5Y cells following MPP+ treatment, a common in vitro Parkinson's disease cell model. Mechanistically, TXNIP ubiquitination through MIB1 E3 ubiquitin ligase regulates NLRP3 inflammasome activation in neuroblasts. In contrast, MPP+ activates the NLPR3 inflammasome through miR590-3p upregulation and direct interference with MIB1-dependent TXNIP ubiquitination. LncZFAS overexpression inhibits this entire pathway through direct interference with miR590-3p, exposing a novel research idea regarding the mechanism of Parkinson's disease. [ABSTRACT FROM AUTHOR]

Introduction: Meningiomas are the most common benign intracranial neoplasms. CNS invasion in meningiomas has been integrated into the 2016 WHO classification of CNS tumors as a stand-alone criterion for atypia. Since then, its prognostic impact has been debated based on contradictory results from retrospective analyses. The aim of the study was to elucidate whether histopathological evidence of CNS invasion is associated with increased proliferative potential. Methods: We have conducted a quantified measurement of the proliferation marker Ki67 and analyzed its association with CNS invasion determined by histology together with other established prognostic markers of progression. Routine, immunohistochemical staining for Ki67 were digitalized and automatic quantification was done using Image J software. Results: Overall, 1718 meningiomas were assessed. Histopathological CNS invasion was seen in 108 cases (6.7%). Uni- and multivariate analysis revealed a significantly higher Ki67 proliferation rate in meningiomas with CNS invasion (p < 0.0001 and p = 0.0098, respectively). Conclusions: Meningiomas with histopathological CNS invasion show a higher proliferative activity. [ABSTRACT FROM AUTHOR]

Despite recent progress in cancer treatment, the prognosis of patients with pancreatic cancer still remains poor. Pancreatic tumors are reported to display high molecular heterogeneity. Elucidating the molecular mechanisms underlying pancreatic cancer progression is essential for improving patient treatment and survival. The overexpression of E3 ubiquitin ligase mind bomb 1 (MIB1) was previously described in pancreatic cancer cells, where it enhanced tumor cell proliferation. However, the role of MIB1 in pancreatic cancer progression remains elusive. In the present study, we confirmed that MIB1 expression is elevated in pancreatic cancer tissues and that high levels of MIB associate with unfavorable prognosis. Overexpression of MIB1 enhanced proliferation and invasion of pancreatic cancer cells both in vitro and in vivo. We further investigated the molecular mechanisms downstream of MIB1 and observed for the first time that MIB1 targets suppressor of tumorigenicity 7 protein (ST7), previously described as suppressor of tumorigenicity, for proteasomal degradation. Furthermore, we found that ST7 suppressed tumor growth by downregulating IQ motif containing GTPase activating protein 1 (IQGAP1) in pancreatic tumor cells. Thus, these data show that MIB1 promotes pancreatic cancer progression by inducing ST7 degradation followed by downregulation of IQGAP1 in pancreatic cancer cells. In conclusion, our research shows that the MIB1/ST7/IQGAP1 axis is essential for pancreatic cancer progression, and MIB1 inhibition may serve as a novel therapeutic strategy in patients with pancreatic cancer. [ABSTRACT FROM AUTHOR]

Abstract Background The dorsal root ganglia (DRG) are a critical component of the peripheral nervous system, and function to relay somatosensory information from the body’s periphery to sensory perception centres within the brain. The DRG are primarily comprised of two cell types, sensory neurons and glia, both of which are neural crest-derived. Notch signalling is known to play an essential role in defining the neuronal or glial fate of bipotent neural crest progenitors that migrate from the dorsal ridge of the neural tube to the sites of the DRG. However, the involvement of Notch ligands in this process and the timing at which neuronal versus glial fate is acquired has remained uncertain. Results We have used tissue specific knockout of the E3 ubiquitin ligase mindbomb1 (Mib1) to remove the function of all Notch ligands in neural crest cells. Wnt1-Cre; Mib1 fl/fl mice exhibit severe DRG defects, including a reduction in glial cells, and neuronal cell death later in development. By comparing formation of sensory neurons and glia with the expression and activation of Notch signalling in these mice, we define a critical period during embryonic development in which early migrating neural crest cells become biased toward neuronal and glial phenotypes. Conclusions We demonstrate active Notch signalling between neural crest progenitors as soon as trunk neural crest cells delaminate from the neural tube and during their early migration toward the site of the DRG. This data brings into question the timing of neuroglial fate specification in the DRG and suggest that it may occur much earlier than originally considered.

Background: The dorsal domain of the neural tube is an excellent model to investigate the generation of complexity during embryonic development. It is a highly dynamic and multifaceted region being first transiently populated by prospective neural crest (NC) cells that sequentially emigrate to generate most of the peripheral nervous system. Subsequently, it becomes the definitive roof plate (RP) of the central nervous system. The RP, in turn, constitutes a patterning center for dorsal interneuron development. The factors underlying establishment of the definitive RP and its segregation from NC and dorsal interneurons are currently unknown. Results: We performed a transcriptome analysis at trunk levels of quail embryos comparing the dorsal neural tube at premigratory NC and RP stages. This unraveled molecular heterogeneity between NC and RP stages, and within the RP itself. By implementing these genes, we asked whether Notch signaling is involved in RP development. First, we observed that Notch is active at the RP-interneuron interface. Furthermore, gain and loss of Notch function in quail and mouse embryos, respectively, revealed no effect on early NC behavior. Constitutive Notch activation caused a local downregulation of RP markers with a concomitant development of dI1 interneurons, as well as an ectopic upregulation of RP markers in the interneuron domain. Reciprocally, in mice lacking Notch activity, both the RP and dI1 interneurons failed to form and this was associated with expansion of the dI2 population. Conclusions: Collectively, our results offer a new resource for defining specific cell types, and provide evidence that Notch is required to establish the definitive RP, and to determine the choice between RP and interneuron fates, but not the segregation of RP from NC. [ABSTRACT FROM AUTHOR]

Left ventricular noncompaction (LVNC) is a structural abnormality of the left ventricle, usually described as an isolated condition, or sometimes associated with other structural cardiac diseases. LVNC is generally asymptomatic, although it may present conduction disorders, arrhythmias, and heart failure. Here, we present the case of a patient who came to our attention with a severe LVNC phenotype associated with advanced AV conduction disorder, and supraventricular and ventricular arrhythmias at young age, in which a novel MIB1, likely pathogenic, variation has been identified.

Background: The Ki‐67 proliferation marker has multiple diagnostic and prognostic applications. Although several clones to the Ki‐67 antigen are commercially available, the MIB1 clone is widely recommended in the surgical pathology literature for neuroendocrine tumors. In our cytopathology practice, we have encountered unexpectedly low MIB1 immunoreactivity in CytoLyt‐fixed cell blocks (CBs). The current study evaluated the impact of fixatives, CB processing, and immunocytochemical (ICC) procedures on Ki‐67 immunoreactivity. Methods: Test CBs were prepared from freshly resected tumors, and multiple variables in the MIB1 ICC procedure were tested, including CytoLyt versus formalin collection media, MIB1 versus other Ki‐67 clones including 30‐9, and other variables. MIB1 versus Ki‐67 30‐9 clones were tested in parallel on CytoLyt‐fixed CBs from clinical samples of small cell lung carcinoma (SCLC). Results: In the test CBs (n = 10), the mean MIB1 labeling index was 10% in CytoLyt versus 47% in formalin (P = .0116), with a mean loss of reactivity in matched CBs of 37% (up to 70%). None of the procedure modifications tested in 223 individual ICC reactions recovered MIB1 reactivity in CytoLyt except for switching to the Ki‐67 30‐9 antibody. In CytoLyt‐fixed SCLC samples (n = 14), the Ki‐67 30‐9 antibody demonstrated expected ranges of reactivity (mean, 83%; range, 60%‐100%), whereas MIB1 demonstrated markedly inhibited labeling (mean, 60%; range, 10%‐95%) (P = .0058). Conclusions: CytoLyt fixation substantially inhibits MIB1 immunoreactivity, whereas the Ki‐67 30‐9 clone is not susceptible to inhibition. Markedly discrepant MIB1 reactivity may present a pitfall in the diagnosis of SCLC and may lead to the incorrect prognostic stratification of other tumor types. For laboratories using CytoLyt, we recommend using the Ki‐67 30‐9 antibody rather than the MIB1 antibody. CytoLyt fixation substantially inhibits MIB1 immunoreactivity and may present a diagnostic pitfall in certain tumors, especially small cell carcinoma. The Ki‐67 30‐9 clone demonstrates adequate performance and should be the clone of choice in laboratories using CytoLyt collection media. [ABSTRACT FROM AUTHOR]

Introduction: Ki‐67 proliferation index (Ki‐67 index) is used to quantify cell proliferation during histopathological assessment of various tumors including glioblastoma (GB). Aim: We aimed to assess correlation between Ki‐67 index and overall survival in patients with GB and determine a cut‐point for Ki‐67 index that predicts for poorer survival. Method: Records of adult patients diagnosed with GB on histopathological specimens at a tertiary cancer center in Sydney between 1 January 2002 and 30 July 2012 were retrieved. Specimens of these patients were examined for quantification of Ki‐67 staining by two independent pathologists. Patient, disease, treatment, and survival data were collected from hospital and cancer care service records. Statistical analysis was performed using proportional hazards models, Kaplan–Meier curves, and the minimum P‐value approach. Result: Of the eligible 71 patients, 58% were males with median age of 58 (range 18–87). Seventy‐three percent of patients were of ECOG performance status 0–1. There was a statistically significant correlation between Ki‐67 index and overall survival. In patients with Ki‐67 > 22% (n = 36), 5‐year survival was approximately 30% compared to 5% in those with Ki‐67 ≤ 22% (n = 35; log‐rank P‐value = 0.04; hazard ratio (HR) = 0.53; 95% confidence intervals (CI), 0.29–0.97). Conclusion: This study demonstrates a positive correlation between Ki‐67 index and overall survival in patients with GB. Percentage staining of Ki‐67 < 22% appears to predict for poorer survival in GB. [ABSTRACT FROM AUTHOR]

Centriolar satellites are non-membrane cytoplasmic granules that deliver proteins to centrosome during centrosome biogenesis and ciliogenesis. Centriolar satellites are highly dynamic during cell cycle or ciliogenesis and how they are regulated remains largely unknown. We report here that sorting nexin 17 (SNX17) regulates the homeostasis of a subset of centriolar satellite proteins including PCM1, CEP131, and OFD1 during serum-starvation-induced ciliogenesis. Mechanistically, SNX17 recruits the deubiquitinating enzyme USP9X to antagonize the mindbomb 1 (MIB1)-induced ubiquitination and degradation of PCM1. SNX17 deficiency leads to enhanced degradation of USP9X as well as PCM1 and disrupts ciliogenesis upon serum starvation. On the other hand, SNX17 is dispensable for the homeostasis of PCM1 and USP9X in serum-containing media. These findings reveal a SNX17/USP9X mediated pathway essential for the homeostasis of centriolar satellites under serum starvation, and provide insight into the mechanism of USP9X in ciliogenesis, which may lead to a better understating of USP9X-deficiency-related human diseases such as X-linked mental retardation and neurodegenerative diseases.

Persistent directional cell migration is involved in animal development and diseases. The small GTPase Rac1 is involved in F-actin and focal adhesion dynamics. Local Rac1 activity is required for persistent directional migration, whereas global, hyperactivated Rac1 enhances random cell migration. Therefore, precise control of Rac1 activity is important for proper directional cell migration. However, the molecular mechanism underlying the regulation of Rac1 activity in persistent directional cell migration is not fully understood. Here, we show that the ubiquitin ligase mind bomb 1 (Mib1) is involved in persistent directional cell migration. We found that knockdown of MIB1 led to an increase in random cell migration in HeLa cells in a wound-closure assay. Furthermore, we explored novel Mib1 substrates for cell migration and found that Mib1 ubiquitinates Ctnnd1. Mib1-mediated ubiquitination of Ctnnd1 K547 attenuated Rac1 activation in cultured cells. In addition, we found that posterior lateral line primordium cells in the zebrafish mib1ta52b mutant showed increased random migration and loss of directional F-actin-based protrusion formation. Knockdown of Ctnnd1 partially rescued posterior lateral line primordium cell migration defects in the mib1ta52b mutant. Taken together, our data suggest that Mib1 plays an important role in cell migration and that persistent directional cell migration is regulated, at least in part, by the Mib1-Ctnnd1-Rac1 pathway. [ABSTRACT FROM AUTHOR]

Summary Notch signaling in neural progenitor cell is triggered by ligands expressed in adjacent cells. To identify the sources of active Notch ligands in the mouse retina, we negatively regulated Notch ligand activity in various neighbors of retinal progenitor cells (RPCs) by eliminating mindbomb E3 ubiquitin protein ligase 1 (Mib1). Mib1-deficient retinal cells failed to induce Notch activation in intra-lineage RPCs, which prematurely differentiated into neurons; however, Mib1 in post-mitotic retinal ganglion cells was not important. Interestingly, Mib1 in the retinal pigment epithelium (RPE) also contributed to Notch activation in adjacent RPCs by supporting the localization of active Notch ligands at RPE-RPC contacts. Combining this RPE-driven Notch signaling and intra-retinal Notch signaling, we propose a model in which one RPC daughter receives extra Notch signals from the RPE to become an RPC, whereas its sister cell receives only a subthreshold level of intra-retinal Notch signal and differentiates into a neuron. [ABSTRACT FROM AUTHOR]

Background: In recent papers, Ki67 labeling index (LI) has been used to classify breast cancer patients into the low and high Ki67LI groups for comparison studies, which showed significant differences in many prognostic factors. It has not been clarified whether image analysis software can be used for calculating LI in breast cancer. In our study, we examined whether Ki67LI in breast cancer calculated using image analysis software correlates with that measured on the basis of visual. Methods: Fifty patients were randomly selected among breast cancer patients who underwent surgical operation from March, 2010 to May, 2010 in our hospital without preoperative chemotherapy. In this study, for the virtual slide system (VSS: VS120-L100, Olympus, Tokyo, Japan), the high-resolution VSs of all the 50 patients were prepared as samples. The image analysis software use for calculating LI was Tissuemorph Digital Pathology (Tissuemorph DP: Visiopharm, Hoersholm, Denmark). The calculated LI was extracted from 3 to 5 views containing hot spots. The LI calculated using Tissuemorph DP was designed as LI/image/T. The digital image of 3 to 5 LI/image/T views was printed out, and on the digital photograph, we counted visually the number of Ki67-immunopositive cells in exactly the same area, and the percentage of Ki67-immunopositive cells was designed as LI/direct. Moreover, a pathologist's assistant (PA) determined the tumor area in the same specimen using VSS and calculated LI using Tissuemorph DP, which was designed as LI/image/PA. The chief pathologist (CP) similarly calculated LI which was designed as LI/image/CP. We evaluated the degree of agreement between different data sets 'LI/image/T and LI/direct' and 'LI/image/T, LI/image/CP, and LI/image/PA' by using interclass correlation coefficient (ICC). Results: The average counts of cells were as follows: LI/direct, 3209.7 ± 1970.4 (SD); LI/image/T, 2601.6 ± 1697.1; LI/image/PA, 2886.5 ± 2027.5; LI/image/CP, 18805.5 ± 22293.4. The values of LI/direct and LI/image/T showed almost perfect agreement as showed by an ICC of 0.885 (95 % CI, 0.806-0.933; p < 0.001). The agreement among three investigators was almost perfect. The obtained ICC was 0.825 (95 % CI, 0.739-0.890; p < 0.001) among the data of LI/image/T, LI/image/CP and LI/image/PA. There were five cases that immunopositivity for Ki67 showed a more than 10 % disagreement between LI/direct and LI/image/T. Conclusion: The merits of calculating Ki67 LI using Tissuemorph DP are as follows. First, the staining intensity of the cells to be counted can be adjusted. Second, the portion of a tumor including 'hot spots' for counting can be chosen. Third, many cancer cells can be counted more rapidly using Tissuemorph DP than by visual observation. However, it is important that pathologist should check and carry out the final decision of the data, when Ki67 LI using Tissuemorph DP is calculated. [ABSTRACT FROM AUTHOR]

Aims: MicroRNA-10 (miR-10) was originally shown to regulate angiogenesis by directly modulating the levels of membranebound fms-related tyrosine kinase 1 (mflt1) and its soluble splice isoform sflt1 post-transcriptionally in zebrafish. Given that flt1 knockdown incompletely rescues the angiogenic phenotypes in miR-10 morphants, flt1 is unlikely to be the only important target of miR-10 in endothelial cells (ECs). It will be interesting to investigate new mechanism responsible for angiogenic defect induced by miR-10 knockdown. Methods and results: Firstly, we demonstrated that miR-10a and miR-10b (miR-10a/10b) were highly enriched in embryonic zebrafish ECs using deep sequencing, Taqman polymerase chain reaction, and in situ hybridisation. Subsequently, we proved that loss of miR-10a/10b impaired blood vessel outgrowth through regulating tip cell behaviours. Mib1 was identified as a potential direct target of miR-10a/10b through in silicon analysis and in vitro luciferase sensor assay. In vivo reporter assay in zebrafish embryos confirmed the binding of miR-10 with 3'-UTR of zebrafish mib1. Furthermore, inhibition of mib1 and Notch signaling rescued the angiogenic defects in miR-10-deficient zebrafish embryos. In addition, we provided evidences that miR-10 regulates human ECs behaviour through targeting Mib1 as well. Conclusion: Taken together, these results indicate that miR-10 regulates the angiogenic behaviour in a Notch-dependent manner by directly targeting mib1. [ABSTRACT FROM AUTHOR]

In the past few years, the monoclonal antibody MIB-1 has been used by researchers in order to retrospectively study paraffin imbibed tumor fragments. The medulloblastoma is the most common malignant central nervous system tumor in childhood. The objectives were: determination of the mean Mib-1 LI value from these patients, as well as the prognostic value of the method.This retrospective study represents an analysis of the cellular proliferation index of posterior fossa medulloblastomas collected from 22 patients at A.C. Camargo Hospital, from January 1990 to December 1999. The histopathological diagnosis was confirmed by H&E and proliferative index (LI) was achived with Mib-1 which detects proliferating cells during G1, G2, S and M phases.The results demostrated that the mean Mib-1 was 30,1%, and ranged from 5,2% to 62,0%.In conclusion, this method has prognostic value, has to be used as routine for patients harboring medulloblastomas and the ones who have PI greater than the mean value found in this study, should be treated aggressively.Nos últimos anos, o anticorpo monoclonal Mib1 tem sido bastante utilizado pelos pesquisadores para estudo retro e prospectivo, pela possibilidade de se obter um índice de proliferação de fragmentos tumorais conservados em parafina. O meduloblastoma é o tumor maligno mais freqüente do sistema nervoso central na infância. Os objetivos do trabalho foram determinar a média IP através do Mib-1 destas neoplasias, e estabelecer seu valor prognóstico. Neste trabalho foi determinado retrospectivamente o índice de proliferação celular de tumores extraídos de 22 pacientes portadores de meduloblastoma da fossa craniana posterior, tratados no Departamento de Neurocirurgia do Hospital A.C. Camargo de S. Paulo, no período de janeiro de 90 a dezembro de 99. O diagnóstico histopatológico de meduloblastoma foi confirmado pela coloração pela hematoxilina e eosina (HE) e o IP foi determinado através do marcador tumoral anticorpo monoclonal Mib1, que detecta as células em proliferação tumoral nas fases G1, G2, S e M do ciclo celular. Os resultados mostraram que a média do IP Mib1 foi de 30,1%, e variou de 5,2% a 62,0%. A conclusão é que este método tem valor prognóstico, deve ser realizado como rotina no diagnóstico anatomopatológico dos pacientes portadores de meduloblastoma e que aqueles com IP Mib1 maior que a média deverão ser submetidos a tratamentos adjuvantes mais agressivos.

Neuroendocrine tumors (NET) are routinely graded and staged to judge prognosis. Proliferation index using MIB1 staining has been introduced to assess grading. There are vivid discussions on cutoff definitions, automated counting, and interobserver variability. However, no data exist regarding interlaboratory reproducibility for low proliferation indices which are of importance to discriminate between G1 and G2 NET. We performed MIB1 staining in three different university hospital-based pathology laboratories on a tissue micro array (TMA) of a well-characterized patient cohort, containing pancreatic NET of 61 patients. To calculate the proliferation index, number of positive tumor nuclei was divided by the total number of tumor nuclei. Labeling index was compared to mitotic counts in whole tissue sections and to clinical outcome. Linear regression analysis, intraclass comparison, and log-rank analysis were performed. Intraclass correlation showed moderate-to-fair agreement. Especially low proliferating tumors were affected by interlaboratory differences. Log-rank analysis was performed for each lab and resulted in three different cutoffs (5.0, 3.0, and 0.5 %). Every calculated cutoff stratified the patient cohort to a significant extent for the underlying stain (p < 0.001, <0.001, and <0.001) but showed no or lesser significance when applied to the other stains. Significant and relevant interlab differences for MIB1 exist. Since the MIB1 proliferation index influences grading, local cutoffs or external standardization should urgently be introduced to achieve reliability and reproducibility. [ABSTRACT FROM AUTHOR]

Plasmacytoma classified into solitary plasmacytoma of bone (SPB) and extramedullary plasmacytoma (EMP) is characterized by infiltrate of plasma cells of diverse maturity and by their monoclonal immunoglobulin products. Both SPB and EMP represent different groups of neoplasm in terms of location, tumor progression, and overall survival rate. There is a need for features that indicate likelihood of myeloma in patients with plasmacytoma without other manifestations. This study was an attempt to study the morphologic patterns of plasmacytoma (SPB and EMP), MIB1 proliferation index, and correlation of these with clinicopathologic features and survival of the patients. The study group comprised of 134 cases of plasmacytoma (88 SPB and 46 EMP) over duration of 8 years and were graded as per Bartl's histologic grading system. Commonest site was vertebral body in SPB (36%) and upper aerodigestive tract in EMP (48%). On serum electrophoresis, overall M band was detected in 41% cases. Both SPB and EMP on histology revealed similar morphologic features. MIB1 proliferation index ranged from less than 1% to 80%. It was slightly higher in EMP in comparison with SPB ( P value = .002). Seventy percent of cases, which progressed to multiple myeloma (MM) showed MIB1 labeling index more than 10%; however, it was not statistically significant in predicting the disease progression. With the median follow-up of 19 months (range, 1-99 months), 10 SPB had disease progression of which 7 converted to MM, and 3 developed EMP, with a median interval of 21 months (range, 8-75 months) for the development of MM and 3 months (range, 3-9 months) for the progression to EMP. Five-year survival for EMP varied by site, with poorest survival in brain/central nervous system EMP as compared with EMP at other sites. To conclude, grade and MIB1 proliferation index help in predicting aggressive course in plasmacytoma. [ABSTRACT FROM AUTHOR]

Aims To compare SP6 and MIB1 antibodies for assessment of Ki67 in primary breast cancer with regard to prognostic value and reproducibility. Methods and results A cohort of 237 premenopausal women with node-negative breast cancer, mainly (87%) not treated with adjuvant systemic therapy, was used. Assessment of Ki67 ( SP6 and MIB1) was performed on tissue microarray by three different investigators. The seventh decile was applied for cut-off. Distant disease-free survival ( DDFS) was chosen as endpoint and the follow-up was restricted to 5 years. Eighty-nine per cent of the samples were classified into the same proliferation group, irrespective of antibody used. For both antibodies, high Ki67 was associated with inferior DDFS in univariable analyses ( SP6: HR 2.5, P = 0.01; and MIB1: HR 2.8, P = 0.004), but failed to reach statistical significance for DDFS in multivariable analyses adjusted for HER2, age, and tumour size ( SP6: HR 2.0, P = 0.074; and MIB1: HR 2.2, P = 0.058). The agreement between different assessors was somewhat higher for MIB1 than for SP6 (κ 0.83-0.88 versus 0.72-0.77). Conclusions SP6 was not superior to MIB1, but the two antibodies were comparable in the assessment of Ki67. Both MIB1 and SP6 could therefore be considered for prognostic use in primary breast cancer. [ABSTRACT FROM AUTHOR]

Integrins are cell surface receptors that mediate cell-cell/extracellular interactions and have shown an association with metastasis or transformation in several cancers. However, the correlation between the clinicopathological status of breast cancer and the altered integrin α4 status is not clear. In this study, we investigated DNA methylation of integrin α4 in breast cancer. We retrieved 351 cases of surgically resected breast cancer (290 invasive carcinoma and 61 intraductal carcinoma). Methylation-specific polymerase chain reaction was performed to determine integrin α4 methylation status. Integrin α4 methylation was frequently observed in breast cancer specimens (145/351 cases, 41.3 %). In addition, DNA methylation of integrin α4 showed statistical correlation with HER2 positivity and higher histologic grade ( p = 0.001, 0.008 in ductal carcinoma in situ and 0.003 in invasive ductal carcinoma). However, other clinicopathological data such as estrogen receptor, progesterone receptor, metastasis, and TNM status showed no statistical correlation. Moreover, we found that the downregulated expression of integrin α4 in a heavily methylated breast cancer cell line (ZR-75) was restored by treatment with 5-aza-2'deoxycytidine, a DNA methyltransferase inhibitor, implying transcriptional silencing by DNA methylation. We observed that integrin α4 methylation is associated with the histologic grade of tumors and lymph node metastasis. Also, this data supports a previous study that suggested integrin α4 and HER2 are involved in the same signaling pathway. DNA methylation of integrin α4 may be a poor prognostic factor which affects undifferentiated histologic change of breast cancer. [ABSTRACT FROM AUTHOR]

The karyopherin protein family comprises importins and exportins which are nucleocytoplasmic shuttling receptors. Increased levels of karyopherin a2 and chromosome region maintenance protein 1 correlate with a higher WHO grade and a poorer prognosis in patients with infiltrative astrocytomas. The aim of this study was to evaluate representative members of importins and exportins (i.e. karyopherin a2 and chromosome region maintenance protein 1) as novel biomarkers for meningiomas of WHO grades I-III. We semiquantitatively analyzed nuclear expression of karyopherin a2, chromosome region maintenance protein 1 and the MIB1 labeling index using immunohistochemistry in 108 primary (44 meningiomas WHO grade I, 48 meningiomas WHO grade II, 16 meningiomas WHO grade III) and 13 recurrent meningiomas. Statistical analysis was performed using standard techniques. Karyopherin a2 ( p < 0.001) and chromosome region maintenance protein 1 ( p = 0.002) expression correlated significantly with the histological grade. Karyopherin a2 expression correlated with proliferative activity as assessed by the MIB1 index ( p < 0.001). Recurrent tumors expressed significantly higher levels of karyopherin a2 ( p = 0.045) when compared to primary growths. Multivariate analysis of the overall series as well as of patients with atypical meningiomas identified higher karyopherin a2 (≥5 vs. <5 %) and chromosome region maintenance protein 1 (≥60 vs. 60 %) expression as independent predictors of tumor recurrence. Karyopherin a2 and chromosome region maintenance protein 1 expression may have potential as novel biomarkers for meningiomas. [ABSTRACT FROM AUTHOR]

The article discusses a research study on comparing markers of cell proliferation in transplanted canine transmissible venereal tumor (TVT). Twelve tumor specimens were examined through the argyrophilic nucleolar organizing regions (AgNOR) technique or staining with Schorr or immunolabelling for Ki-67 antigen (MIB1). Results showed confinement of MIB1 labeling in the nucleus while the AgNOR index on proliferation had positive correlation to the mitotic index and MIB1 immunolabeling.

Introduction: Although intracranial meningiomas are regarded as benign tumors, some of them behave clinically as malignant tumors. Past reports suggest that MIB 1 and vascular endothelial growth factor (VEGF) in postoperative tumor specimens correlate with the aggressive nature of tumors, but preoperative prediction of such a nature is more useful for therapeutic planning for the tumor. The purpose of this study was to assess the usefulness of preoperative thallium-201 chloride single-photon emission computed tomography (Tl SPECT) to evaluate biological behavior in intracranial meningiomas. Methods: Tl SPECT was performed on 39 patients with intracranial meningioma and Tl uptake indices were calculated. The difference in the Tl uptake index between atypical meningiomas and other pathological types of meningioma was evaluated. Moreover, correlation of Tl uptake indices with the MIB1 labeling index was estimated. Tl uptake indices were also compared between VEGF strongly positive and weakly positive meningiomas. Results: The delayed index of atypical meningioma was significantly higher than that of the other pathological types ( p = 0.036). Significant correlation was found between the Tl uptake index in the delayed image and MIB1 labeling index ( p < 0.0001, R = 0.36). Moreover, VEGF strongly positive meningiomas exhibited a significantly higher Tl uptake index compared to VEGF weakly positive meningiomas in both the early image and the delayed image ( p = 0.029, 0.023, respectively). Conclusions: Tl uptake index may be a possible preoperative surrogate marker of MIB1 and VEGF that is useful in detecting aggressive natures in intracranial meningiomas. [ABSTRACT FROM AUTHOR]

Abstract: Background: Erosive esophagitis is a frequent endoscopic feature in patients with gastro-oesophageal reflux disease. However, most of patients with heartburn/regurgitation have a non-erosive reflux disease. The reason for this heterogeneous impact of gastro-oesophageal reflux disease on oesophageal mucosa is unknown to date. Aim: To evaluate the cell proliferation status of oesophageal epithelium in both healthy normal subjects and patients with gastro-oesophageal reflux disease with or without erosions. Materials and methods: All the subjects underwent endoscopy and biopsies were taken at 5cm from the squamo-columnar junction. Specimens were analysed both at histology and at transmission electron microscopy. Cell proliferation was evaluated by MIB1 immunostaining. Of the 85 subjects were studied, 10 were healthy controls with normal pH-testing and macroscopical, histological and ultrastructural patterns; 37were patients with erosive esophagitis, and 38 patients with non-erosive reflux disease. Results: At histology, of the 37 patients affected by erosive esophagitis, 30 had normal mucosa and 7 showed mild oesophagitis. One patient with non-erosive reflux disease showed signs of oesophagitis at histology. At TEM, all patients with gastro-oesophageal reflux disease had ultrastructural patterns of damage i.e. dilations of intercellular spaces (DIS), and all controls had a normal ultrastructural pattern. The mean (±SD) MIB1-LI values of normal subjects and non-erosive reflux disease and erosive oesophagitis patients were 62.2% (±9.1), 29.7% (±7.2) and 16.2% (±5.2), respectively; there were significant differences among the three groups (p <0.001). Conclusions: Oesophageal mucosa of patients with reflux symptoms presents a decrease in MIB1 immunostaining of 50% and 25% in non-erosive reflux disease and erosive esophagitis patients with respect to normal subjects. [Copyright &y& Elsevier]

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Paracoccidioidomycosis (PCMycosis) is a systemic mycosis frequently found in many regions of Latin America. Microscopically, it is characterised by granulomatous inflammation and pseudoepitheliomatous hyperplasia (PEH). This work describes the proliferation index and p53 expression by immunohistochemistry in PEH of PCMycosis, normal oral mucosa (NOM) and mild oral epithelial dysplasia (ED). Ki67 positive cells were present in the basal and parabasal layers in NOM and PEH, while in ED it was also observed in the spinous layer. Percentage of ki67 positive cells was 7.7, 28.2 and 46.0 in NOM, PEH and ED respectively. p53 was negative in NOM and in PEH it was expressed by few cells in the basal layer of only three cases. However, it was expressed in all cases of ED, in basal and parabasal layers. Although histologically PEH mimics well-differentiated squamous cell carcinoma, its proliferative pattern and p53 expression are more similar to NOM than to dysplasia. These findings, confirm PEH as a reactive process probably associated with the underlying chronic inflammation. [ABSTRACT FROM AUTHOR]

Aims: The differentiation of adrenocortical carcinomas from adenomas may be difficult based on morphology alone. Differential expression of insulin-like growth factor (IGF) II and cyclin-dependent kinase (CDK) 4 has recently been described in these tumours. The aim of this study was to investigate the diagnostic usefulness of these markers immunohistochemically. Methods and results: We examined 22 benign and 17 malignant adrenocortical tumours and compared IGFII and CDK4 expression with known immunohistochemical as well as morphological criteria of malignancy. Thirteen of 17 carcinomas showed immunohistochemical reactivity for IGFII, whereas all adenomas but one were negative. Intense CDK4 expression was detected in 11 of 17 carcinomas but was present in only three of 22 adenomas. The MIB1 index was > 5% in 14 of 16 carcinomas and was < 5% in all adenomas but one. The combination of IGFII immunohistochemistry with MIB1 index led to high sensitivity and specificity in detecting adrenocortical carcinomas. Conclusions: IGFII and MIB1 are helpful immunohistochemical markers to predict malignancy in adrenocortical neoplasms. These markers can be used in addition to clinical, gross and morphological features to establish a diagnosis in difficult cases. [ABSTRACT FROM AUTHOR]

At least 25 families with camptodactyly-arthropathy-coxa vara-pericarditis (CACP syndrome) have been reported, with descriptions of a distinctive synovial pathology based largely on light microscopy. Although described as “proliferative,” with numerous multinucleated giant cells, the natures of proliferating cells and giant cells have not been determined. To clarify the pathogenesis of this disorder, we studied 3 patients who had CACP syndrome and underwent synovial biopsy. Cells in the biopsies were studied by immunohistochemistry and electron microscopy. Giant cells were identified as macrophage in origin based on CD68 expression and electron microscopic features of macrophages. Most cells in the synovium were CD68 positive, in keeping with macrophages. The degree of proliferation in synovial biopsies was estimated by MIB1 immunostaining, which showed that up to 30% of cells were cycling compared with fewer than 10% in control synovial biopsies. None of the giant cells was cycling. By double immunostaining, proliferating cells were determined to be fibroblastic synoviocytes rather than macrophages. Thus the proliferative synovitis in this CACP syndrome can be more accurately thought of as hypercellularity by infiltrating macrophages with a contribution by proliferating fibroblastic synoviocytes. The synoviocyte proliferation is likely a response to the underlying genetic mutations involving the proteoglycan-4 (orCACP) gene. The encoded protein normally acts as a lubricant and possibly controls cell proliferation. Loss of one or another of these functions may be a possible mechanism that leads to synoviocyte proliferation in this disease, but the exact pathophysiology leading to this change requires further study. [ABSTRACT FROM AUTHOR]

: ObjectiveImmunohistochemical analysis of MIB1, p53, estrogen, and progesterone receptors can provide prognostic information in endometrial adenocarcinoma. Since predictors of recurrence for low-grade endometrial stromal sarcoma (LESS) are still unknown, a battery of immunostains was performed to find markers, which might be useful to predict prognosis.: MethodsEleven patients with an average age of 43.8 years (range 27–76) were identified with stage I LESS. Immunostains, including MIB1, p53, ER, and PR, were evaluated by two pathologists, independently.: ResultsAll tumors were positive for ER and PR; 1/11 was positive for p53; MIB1 ranged from 0 to 20% positive tumor nuclei. Mitotic counts ranged from 0 to 7/10 hpf. Two patients developed recurrences. One had a pelvic recurrence 7 years after diagnosis. This tumor had a mitotic count of 1/10 hpf, MIB1 expression in 10% of nuclei, and focal p53 expression. A second patient developed pulmonary metastases 10.8 years after diagnosis; the tumor showed a mitotic count of 7/10 hpf and MIB1 expression in 20% of nuclei, but was negative for p53. There was a significant difference in MIB1 reactivity scores between patients who did or did not develop recurrence (P = 0.0303). A marginally significant association was detected between MIB1 (P = 0.0896) or p53 (P = 0.0833) positivity and length of recurrence-free survival.: ConclusionAlthough MIB1 and p53 appear to be useful prognostic markers, a larger study would be necessary to confirm their validity. [Copyright &y& Elsevier]

Aims: Preinvasive endocervical glandular lesions, termed cervical glandular intraepithelial neoplasia, are increasing in incidence. The distinction of cervical glandular intraepithelial neoplasia from benign mimics, especially tubo-endometrial metaplasia, endometriosis and microglandular hyperplasia, can be difficult. This study investigates the value of immunohistochemical staining with MIB1, bcl2 and p16 in the distinction of cervical glandular intraepithelial neoplasia from these benign mimics. Methods and results: Immunohistochemical staining using the monoclonal antibodies MIB1, bcl2 and p16 was performed on cases of cervical glandular intraepithelial neoplasia (n =21), tubo-endometrial metaplasia (n =13), endometriosis (n =7) and microglandular hyperplasia (n =14). With tubo-endometrial metaplasia and microglandular hyperplasia staining with MIB1 was either negative or involved <10% of cells, while with cervical glandular intraepithelial neoplasia the majority of cases (86%) exhibited >10% positive cells. Two cases of endometriosis exhibited a MIB1 index of 10–30% while in the other cases <10% cells stained. With bcl2 the cells of microglandular hyperplasia were negative although there was staining of associated reserve cells in 43% of cases. All cases of tubo-endometrial metaplasia except one and all cases of endometriosis stained diffusely positive with bcl2. Cases of cervical glandular intraepithelial neoplasia were negative or exhibited focal staining. With p16 all cases of cervical glandular intraepithelial neoplasia exhibited diffuse strong positivity, generally involving 100% of cells, while all cases of microglandular hyperplasia were negative. Sixty-two percent of cases of tubo-endometrial metaplasia showed focal positivity, the remainder being negative. Cases of tubo-endometrial metaplasia were never diffusely positive with p16. In three cases of endometriosis there was staining of >50% of cells while the other cases were either focally positive or negative. Conclusions: A panel of antibodies, comprising MIB1, bcl2 and p16, is a useful adjunct to histology in distinguishing cervical glandular intraepithelial neoplasia from tubo-endometrial metaplasia, endometriosis and microglandular hyperplasia. Cases of cervical glandular intraepithelial neoplasia are diffusely positive for p16 and generally exhibit a high proliferation index with MIB1, while bcl2 is negative or, at most, focally positive. Tubo-endometrial metaplasia and endometriosis are characterized by strong diffuse positivity with bcl2 and a low proliferation index with MIB1 (although occasional cases of endometriosis show moderate proliferative activity). p16 is negative or exhibits focal positivity in tubo-endometrial metaplasia but in endometriosis there may be quite widespread positivity. Microglandular hyperplasia shows a low proliferation index with MIB1 and is negative for bcl2 and p16. [ABSTRACT FROM AUTHOR]

Pre-treatment oestrogen receptor (ER) expression in breast cancer predicts for rate of response to endocrine therapy but not for the quality or duration of response (DofR). ER is known to be down-regulated by anti-oestrogens. This study has tested the hypothesis that the degree of down-regulation of ER and the ER-regulated marker bcl-2 are associated with the quality and duration of tamoxifen response. 80 patients with ER+ve breast cancer (H-score ≥10) receiving primary tamoxifen ( n=51 Stage I–II elderl; n=29 Stage III) underwent sequential tumour biopsies for immunocytochemical assessment of ER, bcl-2 and the proliferation marker MIB1. Median follow-up is 45 months. By 6-months on therapy three patients had attained complete response (CR), 27 partial response (PR); 44 static disease (SD) and six progression (PD) by UICC criteria. Greater decrease in ER and bcl-2 H-score from pre-treatment to 6 weeks ( p=0.035, p=0.037) and ER and bcl-2 H-score from pre-treatment to 6 months ( p=0.058, p=0.036) were significantly associated with better quality of response (CR/PR vs SD/PD). Greater 6-week and 6-month reduction in bcl-2 H-score ( p=0.041, p=0.036) and 6-week reduction in MIB1 ( p=0.013) were significantly correlated with longer DofR. This study demonstrates that greater down-regulation of ER and the ER-regulated protein bcl-2 on primary tamoxifen are significantly associated with a better quality of response and bcl-2 and the proliferation marker MIB1 a longer duration of response in ER+ve breast cancer. [ABSTRACT FROM AUTHOR]

Objectives Recently published data suggest a prognostic value of immunohistochemical proliferation markers for limited laryngeal carcinoma. Previous studies have reported contrasting findings on this issue. In this context, different treatment modalities may be responsible for contradictory findings. To study the relationship between proliferative activity-expressed by the immunohistochemical labeling index of proliferation-associated markers Ki-67 (MIB1), Lewis-X (LeuM1), and proliferating cell nuclear antigen (PCNA) and by p53 status-and treatment failure in a matched-pair study on recurrent and nonrecurrent T1 and T2 glottic carcinoma having received primary transoral laser surgery. Methods Twenty-one patients with tumor recurrence were randomly selected and matched with 26 patients with nonrecurrent disease regarding histopathological grading and age. MIB1 staining was used to determine the Ki-67 labeling inde-, and LeuM1 staining for detecting the Lewis-X antigen; immunohistochemistry determined the p53 status and PCNA labeling inde-. Results The Ki-67 labeling index was significantly ( P = .001) higher in tumors from patients who had treatment failure (mean = 20.02%) than in patients who did not fail treatment ('nonfailures') (mean = 9.95%). Carcinoma with a Ki-67 (MIB1) labeling index above the median (15%) of the general study population showed a mean time to relapse of 23 months (n = 21), compared with 50 months for cases (n = 26) below the median ( P = .016). PCNA labeling index correlated less impressively with tumor recurrence (mean = 28.59% for treatment failures, mean = 21.75% for nonfailures, P = .022). Positive detection of the Lewis-X antigen was significantly associated with recurrence ( P = .015) and time to relapse ( P = .006). Status of p53 was not a significant prognostic factor. Conclusion The Ki-67 (MIB1) labeling index may be associated with early relapse of limited laryngeal carcinoma treated with transoral laser surgery. Since the prognostic relevance of Ki-67 seems to be different for radiological and surgical concepts of treatment, Ki-67 might become useful as criterion of therapy selection. The Lewis-X antigen, for the first time used on laryngeal carcinoma, seems to be a strong prognostic marker deserving further investigations. [ABSTRACT FROM AUTHOR]

Northeast Thailand has a very high incidence of intrahepatic cholangiocarcinoma (ICC), which is closely linked to infestation by the liver fluke, whereas the etiology of ICC in Japan remains to be clarified. This study compared the clinicopathological features, the expression of p53 and c-erbB-2 proteins, and the proliferative activity of ICC in 19 Thai and 23 Japanese patients with ICC who were treated by hepatic resection. The average age of the Thai patients (55.8 years) was lower than that of the Japanese (61.3 years). All Thai patients presented with symptoms, whereas 8 Japanese patients were asymptomatic. There were no significant differences in preoperative liver function test values. Tumors were less likely to be located in the right lobe in the Japanese (34.8%) than in the Thai patients (63.2%). Peribiliary fibrosis and adenomatous hyperplasia in noncancerous hepatic tissues were much more frequently found in the Thai patients ( P = 0.0010; P < 0.0001). No significant differences in the expression of p53 protein or c-erbB-2 protein were found between the two series of patients, but proliferative activity, evaluated on the basis of mean MIB1 labeling index, was significantly higher in the Thai patients ( P < 0.001). The present study suggested a higher proliferative activity of ICC in Thai patients than in Japanese patients. [ABSTRACT FROM AUTHOR]