Self-Supervised Learning Reveals Clinically Relevant Histomorphological Patterns for Therapeutic Strategies in Colon Cancer.

Self-supervised learning (SSL) automates the extraction and interpretation of histopathology features on unannotated hematoxylin-and-eosin-stained whole-slide images (WSIs). We trained an SSL Barlow Twins-encoder on 435 TCGA colon adenocarcinoma WSIs to extract features from small image patches. Leiden community detection then grouped tiles into histomorphological phenotype clusters (HPCs). HPC reproducibility and predictive ability for overall survival was confirmed in an independent clinical trial cohort (N=1213 WSIs). This unbiased atlas resulted in 47 HPCs displaying unique and sharing clinically significant histomorphological traits, highlighting tissue type, quantity, and architecture, especially in the context of tumor stroma. Through in-depth analysis of these HPCs, including immune landscape and gene set enrichment analysis, and association to clinical outcomes, we shed light on the factors influencing survival and responses to treatments like standard adjuvant chemotherapy and experimental therapies. Further exploration of HPCs may unveil new insights and aid decision-making and personalized treatments for colon cancer patients.

Clonal Relationship and Mutation Analysis in Lymphoplasmacytic Lymphoma/Waldenström Macroglobulinemia Associated With Diffuse Large B-cell Lymphoma.

Patients with lymphoplasmacytic lymphoma/Waldenström macroglobulinemia (LPL/WM) occasionally develop diffuse large B-cell lymphoma (DLBCL). This mostly results from LPL/WM transformation, although clonally unrelated DLBCL can also arise. LPL/WM is characterized by activating MYD88L265P (>95%) and CXCR4 mutations (~30%), but the genetic drivers of transformation remain to be identified. Here, in thirteen LPL/WM patients who developed DLBCL, the clonal relationship of LPL and DLBCL together with mutations contributing to transformation were investigated. In 2 LPL/WM patients (15%), high-throughput sequencing of immunoglobulin gene rearrangements showed evidence of >1 clonal B-cell population in LPL tissue biopsies. In the majority of LPL/WM patients, DLBCL presentations were clonally related to the dominant clone in LPL, providing evidence of transformation. However, in 3 patients (23%), DLBCL was clonally unrelated to the major malignant B-cell clone in LPL, of which 2 patients developed de novo DLBCL. In this study cohort, LPL displayed MYD88L265P mutation in 8 out of eleven patients analyzed (73%), while CXCR4 mutations were observed in 6 cases (55%). MYD88WT LPL biopsies present in 3 patients (27%) were characterized by CD79B and TNFAIP3 mutations. Upon transformation, DLBCL acquired novel mutations targeting BTG1, BTG2, CD79B, CARD11, TP53, and PIM1. Together, we demonstrate variable clonal B-cell dynamics in LPL/WM patients developing DLBCL, and the occurrence of clonally unrelated DLBCL in about one-quarter of LPL/WM patients. Moreover, we identified commonly mutated genes upon DLBCL transformation, which together with preserved mutations already present in LPL characterize the mutational landscape of DLBCL occurrences in LPL/WM patients.

A significant proportion of classic Hodgkin lymphoma recurrences represents clonally unrelated second primary lymphoma.

Despite high cure rates in classic Hodgkin lymphoma (cHL), relapses are observed. Whether relapsed cHL represents second primary lymphoma or an underlying T-cell lymphoma (TCL) mimicking cHL is underinvestigated. To analyze the nature of cHL recurrences, in-depth clonality testing of immunoglobulin (Ig) and T-cell receptor (TCR) rearrangements was performed in paired cHL diagnoses and recurrences among 60 patients, supported by targeted mutation analysis of lymphoma-associated genes. Clonal Ig rearrangements were detected by next-generation sequencing (NGS) in 69 of 120 (58%) diagnoses and recurrence samples. The clonal relationship could be established in 34 cases, identifying clonally related relapsed cHL in 24 of 34 patients (71%). Clonally unrelated cHL was observed in 10 of 34 patients (29%) as determined by IG-NGS clonality assessment and confirmed by the identification of predominantly mutually exclusive gene mutations in the paired cHL samples. In recurrences of >2 years, ∼60% of patients with cHL for whom the clonal relationship could be established showed a second primary cHL. Clonal TCR gene rearrangements were identified in 14 of 125 samples (11%), and TCL-associated gene mutations were detected in 7 of 14 samples. Retrospective pathology review with integration of the molecular findings were consistent with an underlying TCL in 5 patients aged >50 years. This study shows that cHL recurrences, especially after 2 years, sometimes represent a new primary cHL or TCL mimicking cHL, as uncovered by NGS-based Ig/TCR clonality testing and gene mutation analysis. Given the significant therapeutic consequences, molecular testing of a presumed relapse in cHL is crucial for subsequent appropriate treatment strategies adapted to the specific lymphoma presentation.

Deep learning based tumor-stroma ratio scoring in colon cancer correlates with microscopic assessment.

Background: The amount of stroma within the primary tumor is a prognostic parameter for colon cancer patients. This phenomenon can be assessed using the tumor-stroma ratio (TSR), which classifies tumors in stroma-low (≤50% stroma) and stroma-high (>50% stroma). Although the reproducibility for TSR determination is good, improvement might be expected from automation. The aim of this study was to investigate whether the scoring of the TSR in a semi- and fully automated method using deep learning algorithms is feasible. Methods: A series of 75 colon cancer slides were selected from a trial series of the UNITED study. For the standard determination of the TSR, 3 observers scored the histological slides. Next, the slides were digitized, color normalized, and the stroma percentages were scored using semi- and fully automated deep learning algorithms. Correlations were determined using intraclass correlation coefficients (ICCs) and Spearman rank correlations. Results: 37 (49%) cases were classified as stroma-low and 38 (51%) as stroma-high by visual estimation. A high level of concordance between the 3 observers was reached, with ICCs of 0.91, 0.89, and 0.94 (all P < .001). Between visual and semi-automated assessment the ICC was 0.78 (95% CI 0.23-0.91, P-value 0.005), with a Spearman correlation of 0.88 (P < .001). Spearman correlation coefficients above 0.70 (N=3) were observed for visual estimation versus the fully automated scoring procedures. Conclusion: Good correlations were observed between standard visual TSR determination and semi- and fully automated TSR scores. At this point, visual examination has the highest observer agreement, but semi-automated scoring could be helpful to support pathologists.

Detection of Second Primary Lymphoma in Late Diffuse Large B-cell Lymphoma Recurrences.

Approximately one-third of patients with diffuse large B-cell lymphoma (DLBCL) relapse and often require salvage chemotherapy followed by autologous stem cell transplantation. In most cases, the clonal relationship between the first diagnosis and subsequent relapse is not assessed, thereby potentially missing the identification of second primary lymphoma. In this study, the clonal relationship of 59 paired DLBCL diagnoses and recurrences was established by next-generation sequencing-based detection of immunoglobulin gene rearrangements. Among 50 patients with interpretable results, 43 patients (86%) developed clonally related relapsed disease. This was observed in 100% of early recurrences (<2 years), 80% of the recurrences with an interval between 2 and 5 years, and 73% of late recurrences (≥5 years). On the other hand, 7 (14%) out of 50 patients displayed different dominant clonotypes in primary DLBCL and clinical recurrences, confirming the occurrence of second primary DLBCL; 37% of DLBCL recurrences that occurred ≥4 years after diagnosis were shown to be second primary lymphomas. The clonally unrelated cases were Epstein-Barr virus positive in 43% of the cases, whereas this was only 5% in the relapsed DLBCL cases. In conclusion, next-generation sequencing-based clonality testing in late recurrences should be considered in routine diagnostics to distinguish relapse from second primary lymphoma, as this latter group of patients with DLBCL may benefit from less-intensive treatment strategies.

Correlation of Immunological and Histopathological Features with Gene Expression-Based Classifiers in Colon Cancer Patients.

The purpose of this study was to evaluate the association between four distinct histopathological features: (1) tumor infiltrating lymphocytes, (2) mucinous differentiation, (3) tumor-stroma ratio, plus (4) tumor budding and two gene expression-based classifiers­(1) consensus molecular subtypes (CMS) plus (2) colorectal cancer intrinsic subtypes (CRIS). All four histopathological features were retrospectively scored on hematoxylin and eosin sections of the most invasive part of the primary tumor in 218 stage II and III colon cancer patients from two independent cohorts (AMC-AJCC-90 and AC-ICAM). RNA-based CMS and CRIS assignments were independently obtained for all patients. Contingency tables were constructed and a χ2 test was used to test for statistical significance. Odds ratios with 95% confidence intervals were calculated. The presence of tumor infiltrating lymphocytes and a mucinous phenotype (>50% mucinous surface area) were strongly correlated with CMS1 (p < 0.001 and p = 0.008) and CRIS-A (p = 0.006 and p < 0.001). The presence of mucus (≥ 10%) was associated with CMS3: mucus was present in 64.1% of all CMS3 tumors (p < 0.001). Although a clear association between tumor-stroma ratio and CMS4 was established in this study (p = 0.006), still 32 out of 61 (52.5%) CMS4 tumors were scored as stroma-low, indicating that CMS4 tumors cannot be identified solely based on stromal content. Higher budding counts were seen in CMS4 and CRIS-B tumors (p = 0.045 and p = 0.046). No other associations of the measured parameters were seen for any of the other CRIS subtypes. Our analysis revealed clear associations between histopathologic features and CMS or CRIS subtypes. However, identification of distinct molecular subtypes solely based on histopathology proved to be infeasible. Combining both molecular and morphologic features could potentially improve patient stratification.

Targeting histone methylation to reprogram the transcriptional state that drives survival of drug-tolerant myeloid leukemia persisters.

Although chemotherapy induces complete remission in the majority of acute myeloid leukemia (AML) patients, many face a relapse. This relapse is caused by survival of chemotherapy-resistant leukemia (stem) cells (measurable residual disease; MRD). Here, we demonstrate that the anthracycline doxorubicin epigenetically reprograms leukemia cells by inducing histone 3 lysine 27 (H3K27) and H3K4 tri-methylation. Within a doxorubicin-sensitive leukemia cell population, we identified a subpopulation of reversible anthracycline-tolerant cells (ATCs) with leukemic stem cell (LSC) features lacking doxorubicin-induced H3K27me3 or H3K4me3 upregulation. These ATCs have a distinct transcriptional landscape than the leukemia bulk and could be eradicated by KDM6 inhibition. In primary AML, reprogramming the transcriptional state by targeting KDM6 reduced MRD load and survival of LSCs residing within MRD, and enhanced chemotherapy response in vivo. Our results reveal plasticity of anthracycline resistance in AML cells and highlight the potential of transcriptional reprogramming by epigenetic-based therapeutics to target chemotherapy-resistant AML cells.

Optimized Guide RNA Selection Improves Streptococcus pyogenes Cas9 Gene Editing of Human Hematopoietic Stem and Progenitor Cells.

Ribonucleoproteins (RNPs) are frequently applied for therapeutic gene editing as well as fundamental research because the method is fast, viral free, and shows fewest off target effects. We evaluated various parameters to genetically engineer human hematopoietic stem and progenitor cells (HSPCs) using Streptococcus pyogenes Cas9 (spCas9) RNPs, and achieve gene editing efficiencies up to 80%. We find that guide RNA (gRNA) design is critical to achieve high gene editing efficiencies. However, finding effective gRNAs for HSPCs can be challenging, while the contribution of numerous in silico models is unclear. By screening more than 120 gRNAs, our data demonstrate that in silico gRNA prediction models are ineffective. In this study, we established a time- and cost-efficient in vitro transcribed gRNA screening model in K562 cells that predicts effective gRNAs for HSPCs. RNP based screening thus outperforms in silico modeling and we report that gene editing is equally efficient in distinct CD34+ HSPC subpopulations. Furthermore, no effects on cell proliferation, differentiation, or in vitro hematopoietic lineage commitment were observed. Finally, no upregulation of p21 expression was found, suggesting unperturbed HSPC homeostasis.

Novel Approaches in Molecular Characterization of Classical Hodgkin Lymphoma.

Classical Hodgkin lymphoma (cHL) represents a B-cell lymphoproliferative disease characterized by clonal immunoglobulin gene rearrangements and recurrent genomic aberrations in the Hodgkin Reed-Sternberg cells in a reactive inflammatory background. Several methods are available for the molecular analysis of cHL on both tissue and cell-free DNA isolated from blood, which can provide detailed information regarding the clonal composition and genetic alterations that drive lymphoma pathogenesis. Clonality testing involving the detection of immunoglobulin and T cell receptor gene rearrangements, together with mutation analysis, represent valuable tools for cHL diagnostics, especially for patients with an atypical histological or clinical presentation reminiscent of a reactive lesion or another lymphoma subtype. In addition, clonality assessment may establish the clonal relationship of composite or subsequent lymphoma presentations within one patient. During the last few decades, more insight has been obtained on the molecular mechanisms that drive cHL development, including recurrently affected signaling pathways (e.g., NF-κB and JAK/STAT) and immune evasion. We provide an overview of the different approaches to characterize the molecular composition of cHL, and the implementation of these next-generation sequencing-based techniques in research and diagnostic settings.

Mechanical properties of three-dimensionally printed titanium plates used in jaw reconstruction: preliminary study.

The aim of this study was to compare the mechanical properties of three-dimensionally (3D)-printed and conventional surgical plates used for the repair of maxillary or mandibular defects under the same experimental conditions, and to provide experimental evidence for the future application and clinical trial of 3D-printed individualized surgical plates. For the experimental group, two groups of surgical plates with thicknesses of 2.0 mm and 2.5 mm were designed and 3D-printed by electron beam melting, using Ti-6Al-4V as raw material. Conventional commercially available surgical plates with the same thickness were adopted as the control group. A Vickers hardness tester and universal testing machine were used to measure the mechanical properties of the plates (hardness, bending strength, tensile strength, and yield strength). The mechanical properties of 3D-printed surgical plates were significantly better than those of conventional surgical plates of the same thickness (P < 0.001). Comparing the surgical plates of different thickness, the 2.5 mm-thick plates had the highest bending strength in the experimental group (P < 0.001) and the best hardness (P < 0.001), bending strength (P = 0.001), tensile strength (P = 0.001), and yield strength (P = 0.001) in the control group. No statistical difference was found between the two kinds of plates in the experimental group in terms of hardness (P = 0.060), tensile strength (P = 0.096), and yield strength (P = 0.496). The 3D-printed surgical plates have better mechanical properties than the conventional ones.

Clonality assessment and detection of clonal diversity in classic Hodgkin lymphoma by next-generation sequencing of immunoglobulin gene rearrangements.

Clonality analysis in classic Hodgkin lymphoma (cHL) is of added value for correctly diagnosing patients with atypical presentation or histology reminiscent of T cell lymphoma, and for establishing the clonal relationship in patients with recurrent disease. However, such analysis has been hampered by the sparsity of malignant Hodgkin and Reed-Sternberg (HRS) cells in a background of reactive immune cells. Recently, the EuroClonality-NGS Working Group developed a novel next-generation sequencing (NGS)-based assay and bioinformatics platform (ARResT/Interrogate) to detect immunoglobulin (IG) gene rearrangements for clonality testing in B-cell lymphoproliferations. Here, we demonstrate the improved performance of IG-NGS compared to conventional BIOMED-2/EuroClonality analysis to detect clonal gene rearrangements in 16 well-characterized primary cHL cases within the IG heavy chain (IGH) and kappa light chain (IGK) loci. This was most obvious in formalin-fixed paraffin-embedded (FFPE) tissue specimens, where three times more clonal cases were detected with IG-NGS (9 cases) compared to BIOMED-2 (3 cases). In total, almost four times more clonal rearrangements were detected in FFPE with IG-NGS (N = 23) as compared to BIOMED-2/EuroClonality (N = 6) as judged on identical IGH and IGK targets. The same clonal rearrangements were also identified in paired fresh frozen cHL samples. To validate the neoplastic origin of the detected clonotypes, IG-NGS clonality analysis was performed on isolated HRS cells, demonstrating identical clonotypes as detected in cHL whole-tissue specimens. Interestingly, IG-NGS and HRS single-cell analysis after DEPArray™ digital sorting revealed rearrangement patterns and copy number variation profiles indicating clonal diversity and intratumoral heterogeneity in cHL. Our data demonstrate improved performance of NGS-based detection of IG gene rearrangements in cHL whole-tissue specimens, providing a sensitive molecular diagnostic assay for clonality assessment in Hodgkin lymphoma.

Tumour-stroma ratio outperforms tumour budding as biomarker in colon cancer: a cohort study.

The tumour-stroma ratio (TSR) and tumour budding (TB) are two high-risk factors with potential to be implemented in the next TNM classification. The aim of the current study was to evaluate the practical application of the two biomarkers based on reproducibility, independency and prognostic value. Patients diagnosed with stage II or III colon cancer who underwent surgery between 2005 and 2016 were included. Both TSR and TB were scored on haematoxylin and eosin-stained tissue sections. The TSR, based on the relative amount of stroma, was scored in increments of 10%. TB was scored following the consensus guidelines; a bud was defined as ≤ 4 tumour cells. For analysis, three categories were used. Cohen's kappa was used for reproducibility. The prognostic value was determined with survival analysis. In total, 246 patients were included. The TSR distribution was N = 137 (56%) stroma-low and N = 109 (44%) stroma-high. The TB distribution was TB-low N = 194 (79%), TB-intermediate N = 35 (14%) and TB-high N = 17 (7%). The reproducibility of the TSR was good (interobserver agreement kappa = 0.83 and intraobserver agreement kappa = 0.82), whereas the inter- and intraobserver agreement for scoring TB was moderate (kappa 0.47 and 0.45, respectively). The survival analysis showed an independent prognostic value for disease-free survival for TSR (HR 1.57; 95% CI 1.01-2.44; p = 0.048) and for TB-high (HR 2.01; 95% CI 1.02-3.96; p = 0.043). Based on current results, we suggest the TSR is a more reliable parameter in daily practice due to better reproducibility and independent prognostic value for disease-free survival.

Rapid, Sensitive, and Specific Severe Acute Respiratory Syndrome Coronavirus 2 Detection: A Multicenter Comparison Between Standard Quantitative Reverse-Transcriptase Polymerase Chain Reaction and CRISPR-Based DETECTR.

BACKGROUND: Recent advances in CRISPR-based diagnostics suggest that DETECTR, a combination of reverse-transcriptase loop-mediated isothermal amplification (RT-LAMP) and subsequent Cas12 bystander nuclease activation by amplicon-targeting ribonucleoprotein complexes, could be a faster and cheaper alternative to quantitative reverse-transcription polymerase chain reaction (qRT-PCR) without sacrificing sensitivity and/or specificity. METHODS: In this study, we compare DETECTR with qRT-PCR to diagnose coronavirus disease 2019 on 378 patient samples. Patient sample dilution assays suggest a higher analytical sensitivity of DETECTR compared with qRT-PCR; however, this was not confirmed in this large patient cohort, where we report 95% reproducibility between the 2 tests. RESULTS: These data showed that both techniques are equally sensitive in detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) providing additional value of DETECTR to the currently used qRT-PCR platforms. For DETECTR, different guide ribonucleic acids can be used simultaneously to obviate negative results due to mutations in N-gene. Lateral flow strips, suitable as a point-of-care test, showed a 100% correlation to the high-throughput DETECTR assay. More importantly, DETECTR was 100% specific for SARS-CoV-2 relative to other human coronaviruses. CONCLUSIONS: Because there is no need for specialized equipment, DETECTR could be rapidly implemented as a complementary technically independent approach to qRT-PCR thereby increasing the testing capacity of medical microbiological laboratories and relieving the existent PCR platforms for routine non-SARS-CoV-2 diagnostic testing.

IGFBP7 activates retinoid acid-induced responses in acute myeloid leukemia stem and progenitor cells.

Treatment of acute promyelocytic leukemia (APL) with all-trans retinoic acid (ATRA) in combination with low doses of arsenic trioxide or chemotherapy leads to exceptionally high cure rates (>90%). ATRA forces APL cells into differentiation and cell death. Unfortunately, ATRA-based therapy has not been effective among any other acute myeloid leukemia (AML) subtype, and long-term survival rates remain unacceptably low; only 30% of AML patients survive 5 years after diagnosis. Here, we identified insulin-like growth factor binding protein 7 (IGFBP7) as part of ATRA-induced responses in APL cells. Most importantly, we observed that addition of recombinant human IGFBP7 (rhIGFBP7) increased ATRA-driven responses in a subset of non-APL AML samples: those with high RARA expression. In nonpromyelocytic AML, rhIGFBP7 treatment induced a transcriptional program that sensitized AML cells for ATRA-induced differentiation, cell death, and inhibition of leukemic stem/progenitor cell survival. Furthermore, the engraftment of primary AML in mice was significantly reduced following treatment with the combination of rhIGFBP7 and ATRA. Mechanistically, we showed that the synergism of ATRA and rhIGFBP7 is due, at least in part, to reduction of the transcription factor GFI1. Together, these results suggest a potential clinical utility of IGFBP7 and ATRA combination treatment to eliminate primary AML (leukemic stem/progenitor) cells and reduce relapse in AML patients.

MKL1 deficiency results in a severe neutrophil motility defect due to impaired actin polymerization.

Megakaryoblastic leukemia 1 (MKL1) promotes the regulation of essential cell processes, including actin cytoskeletal dynamics, by coactivating serum response factor. Recently, the first human with MKL1 deficiency, leading to a novel primary immunodeficiency, was identified. We report a second family with 2 siblings with a homozygous frameshift mutation in MKL1. The index case died as an infant from progressive and severe pneumonia caused by Pseudomonas aeruginosa and poor wound healing. The younger sibling was preemptively transplanted shortly after birth. The immunodeficiency was marked by a pronounced actin polymerization defect and a strongly reduced motility and chemotactic response by MKL1-deficient neutrophils. In addition to the lack of MKL1, subsequent proteomic and transcriptomic analyses of patient neutrophils revealed actin and several actin-related proteins to be downregulated, confirming a role for MKL1 as a transcriptional coregulator. Degranulation was enhanced upon suboptimal neutrophil activation, whereas production of reactive oxygen species was normal. Neutrophil adhesion was intact but without proper spreading. The latter could explain the observed failure in firm adherence and transendothelial migration under flow conditions. No apparent defect in phagocytosis or bacterial killing was found. Also, monocyte-derived macrophages showed intact phagocytosis, and lymphocyte counts and proliferative capacity were normal. Nonhematopoietic primary fibroblasts demonstrated defective differentiation into myofibroblasts but normal migration and F-actin content, most likely as a result of compensatory mechanisms of MKL2, which is not expressed in neutrophils. Our findings extend current insight into the severe immune dysfunction in MKL1 deficiency, with cytoskeletal dysfunction and defective extravasation of neutrophils as the most prominent features.

[Study of bond strength of one-piece glass fiber posts-and-cores with flared root canals in vitro].

OBJECTIVE: To compare the effects of three kinds of fiber posts (CAD/CAM one-piece glass fiber posts-and-cores, prefabricated glass fiber post and light curing plastic fiber post) on the bond strength of flared root canals and the effect of thermal cycling on their bond strength. METHODS: Extracted human single teeth (n=90) were endodontically treated and randomly divided into three groups (n = 30 each). The teeth were restored by three kinds of fiber post: CAD/CAM one-piece glass fiber posts-and-cores, prefabricated posts and light curing plastic fiber post. Following post cementation, the specimens were stored in distilled water at 37 °C for 7 days. Half bonded specimens of each group were submitted to thermal cycling (6 000 times, 5 to 55 °C) prior to micro-push-out bond strength test. Fabrication of micro-push-out bond strength test specimens was conducted by precision slicing machine. The micro-push-out bond strength was tested using a universal testing machine, and the failure modes were examined with a stereomicroscope. RESULTS: In CAD/CAM one-piece glass fiber posts-and-cores group, the bond strength of cervical, middle and apical was (9.58±2.67) MPa,(8.62±2.62) MPa,(8.21±2.48) MPa respectively before thermal cycling, and after thermal cycling the bond strength of cervical, middle and apical was (8.14±3.19) MPa,(6.43±2.47) MPa,(6.45±3.20) MPa respectively. In prefabricated posts group, the bonding strength of cervical, middle and apical was (3.89±2.04) MPa,(4.83±1.23) MPa,(4.67±1.86) MPa respectively before thermal cycling, and after thermal cycling the bond strength of cervical, middle and apical was (6.18±1.61) MPa,(5.15±1.94) MPa,(6.39±2.87) MPa respectively. In light curing plastic fiber post group, the bond strength of cervical, middle and apical before thermal cycling was (4.05±2.41) MPa,(1.75±1.70) MPa,(2.60±2.34) MPa respectively, and after thermal cycling the bond strength of cervical, middle and apical was (5.04±2.72) MPa,(1.96±1.70) MPa,(1.34±0.92) MPa respectively. Postal types and root canal regions were found to significantly affect the push-out bond strength. Compared with the other two groups, the one-piece glass fiber posts-and-cores had the highest bonding strength in the cervical, middle and apical. Temperature cycling has no significant effect on the micro push-out bond strength of three kinds of fiber posts. CONCLUSION: One-piece glass fiber posts-and-cores has better bonding strength and excellent bonding performance.

IGFBP7 Induces Differentiation and Loss of Survival of Human Acute Myeloid Leukemia Stem Cells without Affecting Normal Hematopoiesis.

Leukemic stem cells (LSCs) are thought to be the major cause of the recurrence of acute myeloid leukemia (AML) due to their potential for self-renewal. To identify therapeutic strategies targeting LSCs, while sparing healthy hematopoietic stem cells (HSCs), we performed gene expression profiling of LSCs, HSCs, and leukemic progenitors all residing within the same AML bone marrow and identified insulin-like growth factor-binding protein 7 (IGFBP7) as differentially expressed. Low IGFBP7 is a feature of LSCs and is associated with reduced chemotherapy sensitivity. Enhancing IGFBP7 by overexpression or addition of recombinant human IGFBP7 (rhIGFBP7) resulted in differentiation, inhibition of cell survival, and increased chemotherapy sensitivity of primary AML cells. Adding rhIGFBP7 reduced leukemic stem and/or progenitor survival and reversed a stem-like gene signature, but it had no influence on normal hematopoietic stem cell survival. Our data suggest a potential clinical utility of the addition of rhIGFBP7 to current chemotherapy regimens to decrease AML relapse rates.

VEGFC Antibody Therapy Drives Differentiation of AML.

High expression of VEGFC predicts adverse prognosis in acute myeloid leukemia (AML). We therefore explored VEGFC-targeting efficacy as an AML therapy using a VEGFC mAb. VEGFC antibody therapy enforced myelocytic differentiation of clonal CD34+ AML blasts. Treatment of CD34+ AML blasts with the antibody reduced expansion potential by 30% to 50% and enhanced differentiation via FOXO3A suppression and inhibition of MAPK/ERK proliferative signals. VEGFC antibody therapy also accelerated leukemia cell differentiation in a systemic humanized AML mouse model. Collectively, these results define a regulatory function of VEGFC in CD34+ AML cell fate decisions via FOXO3A and serve as a new potential differentiation therapy for patients with AML.Significance: These findings reveal VEGFC targeting as a promising new differentiation therapy in AML. Cancer Res; 78(20); 5940-8. ©2018 AACR.

[Effects of zirconia micron coating on the proliferation and differentiation of osteoblasts].

Objective: To investigate the effects of zirconia micro coating on the proliferation and differentiation of osteoblasts on the surface of zirconia ceramic, and to provide a strategy for zirconia implant surface treatment. Methods: Forty tablets of zirconia ceramic, with the diameter of 15 mm and the thickness of 1.5 mm, were prepared. Then, twenty tablets polished by water sandpaper were taken as the control group, and 20 pieces of the zirconia coating after sintering micron were taken as the experimental group. The micromorphology of the surface of the two groups were observed by scanning electron microscope. The cell morphology after inoculation with MC3T3-E1 of osteoblasts on the surface of the material was investigated for 1, 3, and 5 days by scanning electron microscope. The cell proliferation was detected at 1 and 3 days by methyl thiazolyl tetrazolium. The cell differentiation ability was detected at 3 and 7 days by real-time quantitative PCR. Statistical analysis was conducted by independent sample t test. Results: After coating with zirconia micron particles, pores with the diameter of 1-20 µm could be observed on the surface of the test group of tiles through high temperature sintering. The growth of osteoblasts on the surface of the ceramic chip in the test group and control group exhibited the similar cell morphology. As they were cultured for 1 day, the experimental group exhibited a similar quality of cells as those in the test group (P>0.05). After 3 days' incubation, comparing with the cell quality of the test group (1.067 ± 0.077) (P<0.05), the quality of osteoblasts on the surface of zirconia ceramics coating increased to 1.763±0.165, and the expression of mRNA in alkaline phosphatase (ALP), osteopotin (OPN) and osteocalcin (OCN) also increased with the amount of 1.63±0.28, 1.99±0.41 and 1.60±0.30, respectively, compared with the test group (1.00± 0.00) (P<0.05). Seven days later, the expression of mRNA in Runt-related transcription factor-2 (RNUX2) (1.33±0.19), special AT-rich sequence binding protein-2 (SATB2) (1.64 ± 0.36), as well as alkaline phosphatase (ALP) (1.78±0.40), OPN (2.25±0.36), and OCN (1.88±0.21), showed a remarkably increase compared with the test group (1.00±0.00) (P<0.05). Conclusions: Zirconia micro coating on the surface of zirconia ceramics promoted the proliferation and differentiation of osteoblasts adhered.