Genome-wide CRISPR screens identify PKMYT1 as a therapeutic target in pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease with an overall 5-year survival rate of <12% due to the lack of effective treatments. Novel treatment strategies are urgently needed. Here, PKMYT1 is identified through genome-wide CRISPR screens as a non-mutant, genetic vulnerability of PDAC. Higher PKMYT1 expression levels indicate poor prognosis in PDAC patients. PKMYT1 ablation inhibits tumor growth and proliferation in vitro and in vivo by regulating cell cycle progression and inducing apoptosis. Moreover, pharmacological inhibition of PKMYT1 shows efficacy in multiple PDAC cell models and effectively induces tumor regression without overt toxicity in PDAC cell line-derived xenograft and in more clinically relevant patient-derived xenograft models. Mechanistically, in addition to its canonical function of phosphorylating CDK1, PKMYT1 functions as an oncogene to promote PDAC tumorigenesis by regulating PLK1 expression and phosphorylation. Finally, TP53 function and PRKDC activation are shown to modulate the sensitivity to PKMYT1 inhibition. These results define PKMYT1 dependency in PDAC and identify potential therapeutic strategies for clinical translation.

Effects of Platelet-Rich Plasma-Derived Exosomes on Proliferation and Migration of Tendon Stem/Progenitor Cell.

Objective To investigate the effects of platelet-rich plasma-derived exosomes (PRP-Exos) on the proliferation and migration of tendon stem/progenitor cell (TSPC). Methods PRP-Exos were extracted through the combination of polymer-based precipitation and ultracentrifugation.The morphology,concentration,and particle size of PRP-Exos were identified by transmission electron microscopy and nanoparticle tracking analysis.The expression levels of surface marker proteins on PRP-Exos and platelet membrane glycoproteins were determined by Western blot analysis.Rat TSPC was extracted and cultured,and the expression of surface marker molecules on TSPC was detected using flow cytometry and immunofluorescence staining.The proliferation of TSPC influenced by PRP-Exos was evaluated using CCK-8 assay and EdU assay.The effect of PRP-Exos on the migration of TSPC was evaluated by cell scratch assay and Transwell assay. Results The extracted PRP-Exos exhibit typical saucer-like structures,with a concentration of 4.9×1011 particles/mL,an average particle size of (132.2±56.8) nm,and surface expression of CD9,CD63 and CD41.The extracted TSPC expressed the CD44 protein.PRP-Exos can be taken up by TSPC,and after co-cultured for 48 h,concentrations of 50 and 100 µg/mL of PRP-Exos significantly promoted the proliferation of TSPC (both P<0.001),with no statistical difference between the two concentrations (P=0.283).Additionally,after co-cultured for 24 h,50 µg/mL of PRP-Exos significantly promoted the migration of TSPC (P<0.001). Conclusion Under in vitro culture conditions,PRP-Exos significantly promote the proliferation and migration of rat TSPC.

Ultrasonic shear wave elastography predicts the quality of the residual tendon before the rotator cuff repair.

BACKGROUND AND PURPOSE: Effective evaluation of rotator cuff tear residual tendon quality is the key to surgical repair. However, until now, the evaluation of rotator cuff tissue by ultrasonic shear wave elasticity (SWE) has been controversial. This prospective study analyzed the association between preoperative SWE and arthroscopic residual tendon quality scores. METHODS: The shear wave velocity (SWV) of the deltoid muscle, the supraspinatus tendon, and the supraspinatus muscle were measured in full-thickness rotator cuff tear patients. Tendon quality was scored according to tear size, tendon margin, tendon thickness, and footprint coverage during arthroscopy. The arthroscopic scores were used as the gold standard, and the SWV ratio of tendon and muscle (supraspinatus tendon/deltoid and supraspinatus muscle/deltoid) were calculated and correlated with the arthroscopic scores. RESULT: Eighty-nine patients (129 shoulders) were enrolled, including 89 operation shoulders and 40 control shoulders. In the group of operation shoulders, both the SWV ratios of tendon (SWV-RT) and the SWV ratio of muscle (SWV-RM) were negatively correlated with arthroscopic scores (The correlation coefficient (R) ranged from -0.722 to -0.884 and -0.569 to -0.689). The SWV-RT and SWV-RM of the operation shoulders were significantly lower than that of the control shoulders (p < 0.05). CONCLUSION: SWE could be used to predict the quality of the residual tendon before the rotator cuff repair. SWV of the supraspinatus tendon and muscle was a useful parameter to predict the quality of the residual tendon. CRITICAL RELEVANCE STATEMENT: Measuring the shear wave velocity of the supraspinatus tendon and muscle with SWE is useful for predicting the quality of the residual tendon which is one of the key factors for a successful rotator cuff repair. KEY POINTS: • Evaluating the quality of the residual tendon is important before surgery. • Elasticity measurements were negatively correlated with the arthroscopic score. • SWE is useful for predicting the quality of the residual tendon.

In-situ gelation of fibrin gel encapsulating platelet-rich plasma-derived exosomes promotes rotator cuff healing.

Although platelet-rich plasma-derived exosomes (PRP-Exos) hold significant repair potential, their efficacy in treating rotator cuff tear (RCT) remains unknown. In light of the potential for clinical translation of fibrin gel and PRP-Exos, we evaluated their combined impact on RCT healing and explored suitable gel implantation techniques. In vitro experiments demonstrated that PRP-Exos effectively enhanced key phenotypes changes in tendon stem/progenitor cells. Multi-modality imaging, including conventional ultrasound, shear wave elastography ultrasound, and micro-computed tomography, and histopathological assessments were performed to collectively evaluate the regenerative effects on RCT. The regenerated tendons exhibited a well-ordered structure, while bone and cartilage regeneration were significantly improved. PRP-Exos participated in the healing process of RCT. In-situ gelation of fibrin gel-encapsulated PRP-Exos at the bone-tendon interface during surgery proved to be a feasible gel implantation method that benefits the healing outcome. Comprehensive multi-modality postoperative evaluations were necessary, providing a reliable foundation for post-injury repair.

HucMSCs delay muscle atrophy after peripheral nerve injury through exosomes by repressing muscle-specific ubiquitin ligases.

Cell therapy based on mesenchymal stem cells (MSCs) alleviates muscle atrophy caused by diabetes and aging，however, the impact of human umbilical cord mesenchymal stem cells on muscle atrophy following nerve injury and the underlying mechanisms remain unclear. In this study, we evaluated the therapeutic efficacy of human umbilical cord MSCs (hucMSCs) and hucMSC-derived exosomes (hucMSC-EXOs) for muscle atrophy following nerve injury and identified the underlying molecular mechanisms. Sciatic nerve crush injury in rats and the induction of myotubes in L6 cells were used to determine the ameliorating effect of hucMSCs and hucMSC-EXOs on muscle atrophy. Q-PCR and western blot analyses were used to measure the expression of muscle-specific ubiquitin ligases Fbxo32 (Atrogin1, MAFbx) and Trim63 (MuRF-1). Dual-luciferase reporter gene experiments were conducted to validate the direct binding of miRNAs to their target genes. Local injection of hucMSCs and hucMSC-EXOs mitigated atrophy in the rat gastrocnemius muscle following sciatic nerve crush injury. In vitro, hucMSC-EXOs alleviated atrophy in L6 myotubes. Mechanistic analysis indicated the upregulation of miR-23b-3p levels in L6 myotubes following hucMSC-EXOs treatment. MiR-23b-3p significantly inhibited the expression of its target genes, Fbxo32 and Trim63, and suppressed myotube atrophy. Notably, a miR-23b-3p inhibitor reversed the inhibitory effect of miR-23b-3p on myotube atrophy in vitro. These results suggest that hucMSCs and their exosomes alleviate muscle atrophy following nerve injury. MiR-23b-3p in exosomes secreted by hucMSCs contributes to this mechanism by inhibiting the muscle specific ubiquitination ligases Fbxo32 and Trim63.

Chinese Ultrasound Doctors Association Guideline on Operational Standards for 2-D Shear Wave Elastography Examination of Musculoskeletal Tissues.

The Ultrasound Physician Branch of the Chinese Medical Doctor Association sought to develop evidence-based recommendations on the operational standards for 2-D shear wave elastography examination of musculoskeletal tissues. A consensus panel of 22 Chinese musculoskeletal ultrasound experts reviewed current scientific evidence and proposed a set of 12 recommendations for 13 key issues, including instruments, operating methods, influencing factors and image interpretation. A final consensus was reached through discussion and voting. On the basis of research evidence and expert opinions, the strength of recommendation for each proposition was assessed using a visual analog scale, while further emphasizing the best available evidence during the question-and-answer session. These expert consensus guidelines encourage facilitation of the standardization of clinical practices for collecting and reporting shear wave elastography data.

Tumor cell-intrinsic SETD2 inactivation sensitizes cancer cells to immune checkpoint blockade through the NR2F1-STAT1 pathway.

BACKGROUND: Cancer immunotherapies can induce durable tumor regression, but most patients do not respond. SETD2 mutation has been linked to the efficacy of immune checkpoint inhibitors (ICIs) immunotherapy. The functional importance of the SETD2 inactivation and how to modulate immunotherapy response remains unclear. METHODS: To explore the function of SETD2 in immunotherapy, knockout and subsequent functional experiments were conducted. Bulk RNA-seq, ATAC-seq, Chip-seq and single-cell RNA-seq were performed to dissect the mechanism and explore the immune microenvironment of mouse tumor. Flow cytometry was used to assess cell surface antigen and intratumoral T cell levels. RESULTS: We comprehensively determine the effect of SETD2 inactivation in ICIs therapy and elucidate the mechanistic impact on tumor immunity. Murine syngeneic tumors harboring Setd2 inactivation are sensitive to ICIs. By bulk and single-cell RNA-seq, we further reveal that SETD2 inactivation reprograms intratumoral immune cells and inflames the tumor microenvironment, which is characterized by high infiltration of T cells and enhanced antigen presentation to activate CD8+ T cell-mediated killing. Mechanistically, via an integrated multiomics analysis using ATAC-seq, ChIP-seq and RNA-seq, we demonstrate that SETD2 inactivation reduces NR2F1 transcription by impairing H3K36me3 deposition and chromatin accessibility, which activates the STAT1 signaling pathway to promote chemokines and programmed cell death protein-1 (PD-1) expression and enhance antigen presentation. All these regulatory mechanisms synergistically promote the effects of anti-programmed cell death ligand 1 immunotherapy in Setd2-knockout syngeneic mouse models. The SETD2-NR2F1-STAT1 regulatory axis is conserved in human and murine cancers. Finally, cancer patients harboring SETD2 mutations who received ICIs show increased durable clinical benefits and survival. CONCLUSIONS: These findings provide novel insights into the biology of SETD2 inactivation regulation and reveal a new potential therapeutic biomarker for ICIs immunotherapy in various refractory cancers.

The value of combined ultrasound contrast arthrography and subacromial-subdeltoid bursography for detecting and differentiating the rotator cuff tear subtypes in patients with the uncertain rotator cuff tear.

OBJECTIVES: In this study, ultrasound (US) contrast arthrography and subacromial-subdeltoid bursography with the contrast agent of SonoVue were performed to evaluate their value for detecting and differentiating the rotator cuff tear (RCT) subtypes in patients with the uncertain RCT. METHODS: A total of 102 patients with the clinically suspected RCTs in the orthopedic clinic were prospectively recruited and underwent conventional high-frequency US for the category of undoubted full-thickness RCT, uncertain RCT, and intact rotator cuff. Among these patients, the patients with uncertain RCT underwent the subsequent US contrast arthrography and subacromial-subdeltoid bursography. The arthroscopic findings were used as the gold standard in this study. RESULTS: After the conventional US screening, 62 patients with uncertain RCT underwent the subsequent US contrast arthrography and subacromial-subdeltoid bursography. All the US contrast arthrography and subacromial-subdeltoid bursography were successfully performed and no severe side effects were observed in all the patients. For full-thickness tears, the sensitivity and specificity of the combined US contrast arthrography and subacromial-subdeltoid bursography were 94.7% (CI: 0.72-1.0) and 81.4% (CI: 0.66-0.91), respectively, and for articular-side tears 100% (CI: 0.51-1) and 100% (CI: 0.92-1), respectively, and for the bursal-side tears 84.6% (CI: 0.54-0.97) and 97.9% (CI: 0.88-1.0), respectively. The main inconsistency between the contrast-enhanced US and arthroscopy was that 7 patients with arthroscopic proved concurrent articular- and bursal-side tears were indicated as full-thickness RCTs on contrast-enhanced US. CONCLUSIONS: Combined US contrast arthrography and subacromial-subdeltoid bursography are useful for detecting the RCT subtypes in patients with the uncertain RCTs. CLINICAL RELEVANCE STATEMENT: When conventional high-frequency US has some difficulty in differentiating the full-thickness from partial-thickness RCTs, combined US contrast arthrography and subacromial-subdeltoid bursography could be used to improve the detection accuracy of RCT subtypes. KEY POINTS: • This is the first study by injection of the US contrast agent SonoVue into the shoulder joint cavity and subacromial-subdeltoid bursa for the detection and differentiation of the RCT subtypes among the people with the uncertain RCT by conventional US screening. • The SonoVue was injected into the glenohumeral joint cavity under US guidance to differentiate the full-thickness RCTs from partial-thickness RCTs. • Combined US contrast arthrography and subacromial-subdeltoid bursography are useful for detecting the RCT subtypes in patients with the uncertain RCTs.

Ultrasound-Targeted Microbubble Destruction Assisted Delivery of Platelet-Rich Plasma-Derived Exosomes Promoting Peripheral Nerve Regeneration.

Peripheral nerve injury is prevalent and has a high disability rate in clinical settings. Current therapeutic methods have not achieved satisfactory efficacy, underscoring the need for novel approaches to nerve restoration that remains an active area of research in neuroscience and regenerative medicine. In this study, we isolated platelet-rich plasma-derived exosomes (PRP-exos) and found that they can significantly enhance the proliferation, migration, and secretion of trophic factors by Schwann cells (SCs). In addition, there were marked changes in transcriptional and expression profiles of SCs, particularly via the upregulation of genes related to biological functions involved in nerve regeneration and repair. In the rat model of sciatic nerve crush injury, ultrasound-targeted microbubble destruction (UTMD) enhanced the efficiency of PRP-exos delivery to the injury site. This approach ensured a high concentration of PRP-exos in the injured nerve and improved the therapeutic outcomes. In conclusion, PRP-exos may promote nerve regeneration and repair, and UTMD may increase the effectiveness of targeted PRP-exos delivery to the injured nerve and enhance the therapeutic effect.

Value of Multimodality Ultrasound in Quantitative Evaluation of the Intra-compartmental Pressure and Perfusion Pressure in Acute Compartment Syndrome in a Rabbit Model.

OBJECTIVE: The aim of the work described here was to investigate the feasibility of using multimodality ultrasound in quantitative evaluation of the intra-compartmental pressure (ICP) and perfusion pressure (PP) changes in acute compartment syndrome (ACS). METHODS: Infusion technique was used to increase the ICP of the anterior compartment of 10 rabbits from baseline to 20, 30, 40, 50, 60, 70 and 80 mmHg. The anterior compartment was evaluated with conventional ultrasound, shear wave elastography (SWE) and contrast-enhanced ultrasound (CEUS). The shape of the anterior compartment, shear wave velocity (SWV) of the tibialis anterior (TA) muscle and CEUS parameters of the TA muscle were measured. RESULTS: When the ICP exceeded 30 mmHg, the shape of the anterior compartment did not expand significantly with increasing ICP. There was a strong correlation between the SWV of TA muscle and measured ICP (ρ = 0.927). Arrival time (AT), time to peak (TTP), peak intensity (PI) and area under the curve (AUC) were significantly correlated with PP (AT, ρ = -0.763; TTP, ρ = -0.900; PI, ρ = 0.665; AUC, ρ = 0.706), whereas mean transit time (MTT) was not. CONCLUSION: Multimodality ultrasound can be used to quantitatively evaluate ICP and PP and, thus, could provide more information for the rapid diagnosis and monitoring of ACS.

[Human Platelet-Rich Plasma-Derived Exosomes Promote the Proliferation of Schwann Cells Cultured in Vitro].

Objective To investigate the effect of human platelet-rich plasma-derived exosomes(PRP-exos)on the proliferation of Schwann cell(SC)cultured in vitro. Methods PRP-exos were extracted by polymerization-precipitation combined with ultracentrifugation.The morphology of PRP-exos was observed by transmission electron microscopy,and the concentration and particle size distribution of PRP-exos were determined by nanoparticle tracking analysis.Western blotting was employed to determine the expression of the marker proteins CD63,CD81,and CD9 on exosome surface and the platelet membrane glycoprotein CD41.The SCs of rats were isolated and cultured,and the expression of the SC marker S100ß was detected by immunofluorescence staining.The fluorescently labeled PRP-exos were co-cultured with SCs in vitro for observation of their interaction.EdU assay was employed to detect the effect of PRP-exos on SC proliferation,and CCK-8 assay to detect the effects of PRP-exos at different concentrations(0,10,20,40,80,and 160 µg/ml)on SC proliferation. Results The extracted PRP-exos appeared as uniform saucer-shaped vesicles with the average particle size of(122.8±38.7)nm and the concentration of 3.5×1012 particles/ml.CD63,CD81,CD9,and CD41 were highly expressed on PRP-exos surface(P<0.001,P=0.025,P=0.004,and P=0.032).The isolated SCs expressed S100ß,and PRP-exos could be taken up by SCs.PRP-exos of 40,80,and 160 µg/ml promoted the proliferation of SCs,and that of 40 µg/ml showed the best performance(all P<0.01). Conclusions High concentrations of PRP-exos can be extracted from PRP.PRP-exos can be taken up by SCs and promote the proliferation of SCs cultured in vitro.

Activity of Anlotinib in the Second-Line Therapy of Metastatic Gastrointestinal Stromal Tumors: A Prospective, Multicenter, In Vitro Study.

BACKGROUND: Anlotinib is a multi-target tyrosine kinase inhibitor that can effectively inhibit tumor cell proliferation after receptor kinase activation caused by KIT gene mutation. METHODS: We tested the inhibitory effect of anlotinib in GIST cell lines with different gene mutations and evaluated the efficacy of anlotinib for patients with metastatic GIST after imatinib failure in a multicenter, single-arm, phase II study. RESULTS: In vitro, V654A mutation encoded by KIT exon 13 was intermediately sensitive to anlotinib. Moreover, anlotinib was able to partly suppress the activation loop mutation D820A from exon 17 while another activation loop mutation N822K, also from exon 17, was resistant to anlotinib. From September 2018 to October 2020, 64 patients from 9 Chinese medical centers were enrolled in this study. Seven patients had partial response and 39 patients had stable disease. The median PFS was 8.0 months. There was no statistical significance comparing with PFS of sunitinib second-line therapy at the same period. The most common adverse events related to anlotinib treatment were hypertension, neutropenia, and fatigue. CONCLUSION: Anlotinib showed moderate antitumor activity in drug-resistant GIST cell lines in vitro, and good PFS and better tolerance in second-line therapy study.

Clinical value of three-dimensional ultrasonography in the morphologic evaluation of rotator cuff tear: a prospective study.

OBJECTIVE: To study the clinical value of three-dimensional ultrasonography (3D-US) in the morphological evaluation of rotator cuff tears (RCTs). METHODS: Based on previously published literature, RCT patterns in our study were divided into crescent, L-shaped with the remnant tendon retracted to the anterior rotator cuff (aL-shaped), L-shaped with the remnant tendon retracted to the posterior rotator cuff (pL-shaped), T-shaped (a tear pattern that is a combination of aL-shaped and pL-shaped), U-shaped, and massive type. Two radiologists prospectively assessed the tear patterns using 3D-US as well as magnetic resonance imaging (MRI) and compared these results using arthroscopy to calculate diagnostic accuracy. RESULT: Fifty-two patients (52 shoulders) were enrolled. The overall diagnostic accuracy of 3D-US in evaluating RCT patterns (82.7%, 43/52; 95% CI: 72.1-93.3%) was significantly higher (p = 0.008) than that of the MRI (57.7%, 31/52; 95% CI: 45.8-73.4%). The accuracy of 3D-US was higher than that of MRI for most types of tears (crescent: 95.0% vs. 55.0%, aL-shaped: 83.3% vs. 77.8%, pL-shaped: 50.0% vs. 25.0%, T-shaped: 75.0% vs. 0.0%, and massive type: 80.0% vs. 100.0%). The accuracies of 3D-US with respect to evaluation by the two radiologists were 84.6% (44/52) and 76.9% (40/52), and there was substantial agreement evident (κ = 0.709). The time taken by the two radiologists to reconstruct the 3D-US images and evaluate the tear pattern was < 5 min. CONCLUSION: The 3D-US can be used for the preoperative evaluation of RCT patterns, and thus be useful for the correct selection of the surgical repair technique for RCTs. KEY POINTS: • Few studies have been found exploring the value of 3D-US for the morphological evaluation of RCTs and correlated with the arthroscopic findings. • Based on previous studies on the morphological classification, anterior L shape (aL-shaped), and posterior L shape (pL-shaped) were used for the first time to describe the torn patterns of RCT.

Lipidomics and Transcriptomics Differ Liposarcoma Differentiation Characteristics That Can Be Altered by Pentose Phosphate Pathway Intervention.

Liposarcoma (LPS) is a rare and heterogeneous malignancy of adipocytic origin. Well-differentiated liposarcoma (WDLPS) and dedifferentiated liposarcoma (DDLPS) are two of the most common subtypes, showing similar genetic characterizations but distinct biological behaviors and clinical prognosis. Compared to WDLPS, DDLPS is more aggressive and has the potential of metastasis, as the malignant adipocytic tumor's metabolic changes may have taken place during the tumorigenesis of LPSs. Therefore, to investigate the lipid alterations between the two subtypes, high-resolution liquid chromatography tandem mass spectrometry (LC-MS/MS) based untargeted lipidomic analysis was performed onto LPS tissues from 6 WDLPS and 7 DDLPS patients. The lipidomic analysis showed the upregulated phosphatidylcholines and phosphoethanolamines in DDLPS, and the upregulated triglycerides and diglycerides in WDLPS, which might be due to the uncompleted adipocytic dedifferentiation leading to such tumorigenesis. Such a finding was also confirmed by the similarity comparison of two LPS subtypes to the transcriptome of stromal vascular fraction at different differentiation stages. Transcriptomic analysis also demonstrated that metabolic pathways including the pentose phosphate pathway (PPP) were upregulated in WDLPS compared to DDLPS. Therefore, the cell line LPS853 was treated with the PPP inhibitor 6-aminonicotinamide ex vivo and the proliferation and invasion of LPS853 was significantly promoted by PPP inhibition, suggesting the potential role of PPP in the development and differentiation of LPS. In conclusion, this study described the altered lipid profiles of WDLPS and DDLPS for the first time, revealing the different differentiation stages of the two subtypes and providing a potential metabolic target for LPS treatment.

Peripheral nerve defects repaired with autogenous vein grafts filled with platelet-rich plasma and active nerve microtissues and evaluated by novel multimodal ultrasound techniques.

BACKGROUND: Developing biocompatible nerve conduits that accelerate peripheral nerve regeneration, lengthening and functional recovery remains a challenge. The combined application of nerve microtissues and platelet-rich plasma (PRP) provides abundant Schwann cells (SCs) and various natural growth factors and can compensate for the deficiency of SCs in the nerve bridge, as well as the limitations of applying a single type of growth factor. Multimodal ultrasound evaluation can provide additional information on the stiffness and microvascular flow perfusion of the tissue. This study was designed to investigate the effectiveness of a novel tissue-engineered nerve graft composed of an autogenous vein, nerve microtissues and PRP in reconstructing a 12-mm tibial nerve defect and to explore the value of multimodal ultrasound techniques in evaluating the prognosis of nerve repair. METHODS: In vitro, nerve microtissue activity was first investigated, and the effects on SC proliferation, migration, factor secretion, and axonal regeneration of dorsal root ganglia (DRG) were evaluated by coculture with nerve microtissues and PRP. In vivo, seventy-five rabbits were equally and randomly divided into Hollow, PRP, Micro-T (Microtissues), Micro-T + PRP and Autograft groups. By analysing the neurological function, electrophysiological recovery, and the comparative results of multimodal ultrasound and histological evaluation, we investigated the effect of these new nerve grafts in repairing tibial nerve defects. RESULTS: Our results showed that the combined application of nerve microtissues and PRP could significantly promote the proliferation, secretion and migration of SCs and the regeneration of axons in the early stage. The Micro-T + PRP group and Autograft groups exhibited the best nerve repair 12 weeks postoperatively. In addition, the changes in target tissue stiffness and microvascular perfusion on multimodal ultrasound (shear wave elastography; contrast-enhanced ultrasonography; Angio PlaneWave UltrasenSitive, AngioPLUS) were significantly correlated with the histological results, such as collagen area percentage and VEGF expression, respectively. CONCLUSION: Our novel tissue-engineered nerve graft shows excellent efficacy in repairing 12-mm defects of the tibial nerve in rabbits. Moreover, multimodal ultrasound may provide a clinical reference for prognosis by quantitatively evaluating the stiffness and microvescular flow of nerve grafts and targeted muscles.

Characterization of the genomic landscape in large-scale Chinese patients with pancreatic cancer.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is a malignant tumor with an extremely poor prognosis. Effective targets for anticancer therapy confirmed in PDAC are limited. However, the characteristics of genomics have not been fully elucidated in large-scale patients with PDAC from China. METHODS: We collected both blood and tissue samples from 1080 Chinese patients with pancreatic cancer and retrospectively investigated the genomic landscape using next-generation sequencing (NGS). FINDINGS: We found recurrent somatic mutations in KRAS (83.2%), TP53 (70.6%), CDKN2A (28.8%), SMAD4 (23.0%), ARID1A (12.8%) and CDKN2B (8.9%) in Chinese PDAC patients. Compared with primary pancreatic cancers, more genomic alterations accumulated especially cell cycle regulatory gene variants (45.4% vs 31.6%, P < 0.001) were observed in metastatic tumors. The most common mutation site of KRAS is p.G12D (43.6%) in pancreatic cancer. Patients with KRAS mutations were significantly associated with older age and mutations in the other three driver genes, while KRAS wild-type patients contained more fusion mutations and alternative mechanisms of RTK/Ras/MAPK pathway including a number of clinically targetable mutations. KRAS mutations in Chinese cohort were significantly lower than those in Western cohorts (all P < 0.05). A total of 252 (23.3%) patients with the core DNA damage response (DDR) gene mutations were detected. ATM (n =59, 5.5%) was the most frequent mutant DDR gene in patients with pancreatic cancer from China. Patients with germline DDR gene mutations were younger (P = 0.018), while patients with somatic DDR gene mutations were more likely to accumulate in metastatic lesions (P < 0.001) and had higher TMB levels (P < 0.001). In addition, patients with mutant DDR genes and patients carrying TP53 mutation were observed mutually exclusive (P < 0.001). INTERPRETATION: We demonstrated the real-world genomic characteristics of large-scale patients with pancreatic cancer from China which may have promising implications for further clinical significance and drug development. FUNDING: The funders are listed in the Acknowledgement.

Ultrasound microvasculature imaging with entropy-based radiality super-resolution (ERSR).

Objective:Microvasculature is highly relevant to the occurrence and development of pathologies such as cancer and diabetes. Ultrasound localization microscopy (ULM) has bypassed the diffraction limit and demonstrated its great potential to provide new imaging modality and establish new diagnostic criteria in clinical application. However, sparse microbubble distribution can be a significant bottleneck for improving temporal resolution, even for further clinical translation. Other important challenges for ULM to tackle in clinic also include high microbubble concentration and low frame rate.Approach:As part of the efforts to facilitate clinical translation, this paper focused on the low frame rate and the high microbubble distribution issue and proposed a new super-resolution imaging strategy called entropy-based radiality super-resolution (ERSR). The feasibility of ERSR is validated with simulations, phantom experiment and contrast-enhanced ultrasound scan of rabbit sciatic nerve with clinical accessible ultrasound system.Main results:ERSR can achieve 10 times improvement in spatial resolution compared to conventional ultrasound imaging, higher temporal resolution (∼10 times higher) and contrast-to-noise ratio under high-density microbubbles, compared with ULM under low-density microbubbles.Significance:We conclude ERSR could be a valuable imaging tool with high spatio-temporal resolution for clinical diagnosis and assessment of diseases potentially.

E3 ligase MKRN3 is a tumor suppressor regulating PABPC1 ubiquitination in non-small cell lung cancer.

Central precocious puberty (CPP), largely caused by germline mutations in the MKRN3 gene, has been epidemiologically linked to cancers. MKRN3 is frequently mutated in non-small cell lung cancers (NSCLCs) with five cohorts. Genomic MKRN3 aberrations are significantly enriched in NSCLC samples harboring oncogenic KRAS mutations. Low MKRN3 expression levels correlate with poor patient survival. Reconstitution of MKRN3 in MKRN3-inactivated NSCLC cells directly abrogates in vitro and in vivo tumor growth and proliferation. MKRN3 knockout mice are susceptible to urethane-induced lung cancer, and lung cell-specific knockout of endogenous MKRN3 accelerates NSCLC tumorigenesis in vivo. A mass spectrometry-based proteomics screen identified PABPC1 as a major substrate for MKRN3. The tumor suppressor function of MKRN3 is dependent on its E3 ligase activity, and MKRN3 missense mutations identified in patients substantially compromise MKRN3-mediated PABPC1 ubiquitination. Furthermore, MKRN3 modulates cell proliferation through PABPC1 nonproteolytic ubiquitination and subsequently, PABPC1-mediated global protein synthesis. Our integrated approaches demonstrate that the CPP-associated gene MKRN3 is a tumor suppressor.

MKRN3-mediated ubiquitination of Poly(A)-binding proteins modulates the stability and translation of GNRH1 mRNA in mammalian puberty.

The family of Poly(A)-binding proteins (PABPs) regulates the stability and translation of messenger RNAs (mRNAs). Here we reported that the three members of PABPs, including PABPC1, PABPC3 and PABPC4, were identified as novel substrates for MKRN3, whose deletion or loss-of-function mutations were genetically associated with human central precocious puberty (CPP). MKRN3-mediated ubiquitination was found to attenuate the binding of PABPs to the poly(A) tails of mRNA, which led to shortened poly(A) tail-length of GNRH1 mRNA and compromised the formation of translation initiation complex (TIC). Recently, we have shown that MKRN3 epigenetically regulates the transcription of GNRH1 through conjugating poly-Ub chains onto methyl-DNA bind protein 3 (MBD3). Therefore, MKRN3-mediated ubiquitin signalling could control both transcriptional and post-transcriptional switches of mammalian puberty initiation. While identifying MKRN3 as a novel tissue-specific translational regulator, our work also provided new mechanistic insights into the etiology of MKRN3 dysfunction-associated human CPP.