Inhibition of palmitoyltransferase ZDHHC12 sensitizes ovarian cancer cells to cisplatin through ROS-mediated mechanisms.

Platinum-based therapies have revolutionized the treatment of high-grade serous ovarian cancer (HGSOC). However, high rates of disease recurrence and progression remain a major clinical concern. Impaired mitochondrial function and dysregulated reactive oxygen species (ROS), hallmarks of cancer, hold potential as therapeutic targets for selectively sensitizing cisplatin treatment. Here, we uncover an oncogenic role of the palmitoyltransferase ZDHHC12 in regulating mitochondrial function and ROS homeostasis in HGSOC cells. Analysis of The Cancer Genome Atlas (TCGA) ovarian cancer data revealed significantly elevated ZDHHC12 expression, demonstrating the strongest positive association with ROS pathways among all ZDHHC enzymes. Transcriptomic analysis of independent ovarian cancer datasets and the SNU119 cell model corroborated this association, highlighting a strong link between ZDHHC12 expression and signature pathways involving mitochondrial oxidative metabolism and ROS regulation. Knockdown of ZDHHC12 disrupted this association, leading to increased cellular complexity, ATP levels, mitochondrial activity, and both mitochondrial and cellular ROS. This dysregulation, achieved by the siRNA knockdown of ZDHHC12 or treatment with the general palmitoylation inhibitor 2BP or the fatty acid synthase inhibitor C75, significantly enhanced cisplatin cytotoxicity in 2D and 3D spheroid models of HGSOC through ROS-mediated mechanisms. Markedly, ZDHHC12 inhibition significantly augmented the anti-tumor activity of cisplatin in an ovarian cancer xenograft tumor model, as well as in an ascites-derived organoid line of platinum-resistant ovarian cancer. Our data suggest the potential of ZDHHC12 as a promising target to improve the outcome of HGSOCs in response to platinum-based chemotherapy.

Impact of ovary-intact menopause in a mouse model of heart failure with preserved ejection fraction.

Heart failure with preserved ejection fraction (HFpEF) afflicts over half of all patients with heart failure and is a debilitating and fatal syndrome affecting postmenopausal women more than any other demographic. This bias toward older females calls into question the significance of menopause in the development of HFpEF, but this question has not been probed in detail. In this study, we report the first investigation into the impact of ovary-intact menopause in the context of HFpEF. To replicate the human condition as faithfully as possible, vinylcyclohexene dioxide (VCD) was used to accelerate ovarian failure (AOF) in female mice while leaving the ovaries intact. HFpEF was established with a mouse model that involves two stressors typical in humans: a high-fat diet and hypertension induced from the nitric oxide synthase inhibitor NG-nitro-l-arginine methyl ester (l-NAME). In young female mice, AOF or HFpEF-associated stressors independently induced abnormal myocardial strain indicative of early subclinical systolic and diastolic cardiac dysfunction. HFpEF but not AOF was associated with elevations in systolic blood pressure. Increased myocyte size and reduced myocardial microvascular density were not observed in any group. Also, a broad panel of measurements that included echocardiography, invasive pressure measurements, histology, and serum hormones revealed no interaction between AOF and HFpEF. Interestingly, AOF did evoke a higher density of infiltrating cardiac immune cells in both healthy and HFpEF mice, suggestive of proinflammatory effects. In contrast to young mice, middle-aged "old" mice did not exhibit cardiac dysfunction from estrogen deprivation alone or from HFpEF-related stressors.NEW & NOTEWORTHY This is the first preclinical study to examine the impact of ovary-intact menopause [accelerated ovarian failure (AOF)] on HFpEF. Echocardiography of young female mice revealed early evidence of diastolic and systolic cardiac dysfunction apparent only on strain imaging in HFpEF only, AOF only, or the combination. Surprisingly, AOF did not exacerbate the HFpEF phenotype. Results in middle-aged "old" females also showed no interaction between HFpEF and AOF and, importantly, no cardiovascular impact from HFpEF or AOF.

In Vivo MRI Tracking of Degradable Polyurethane Hydrogel Degradation In Situ Using a Manganese Porphyrin Contrast Agent.

BACKGROUND: A noninvasive method to track implanted biomaterials is desirable for real-time monitoring of material interactions with host tissues and assessment of efficacy and safety. PURPOSE: To explore quantitative in vivo tracking of polyurethane implants using a manganese porphyrin (MnP) contrast agent containing a covalent binding site for pairing to polymers. STUDY TYPE: Prospective, longitudinal. ANIMAL MODEL: Rodent model of dorsal subcutaneous implants (10 female Sprague Dawley rats). FIELD STRENGTH/SEQUENCE: A 3-T; two-dimensional (2D) T1-weighted spin-echo (SE), T2-weighted turbo SE, three-dimensional (3D) spoiled gradient-echo T1 mapping with variable flip angles. ASSESSMENT: A new MnP-vinyl contrast agent to covalently label polyurethane hydrogels was synthesized and chemically characterized. Stability of binding was assessed in vitro. MRI was performed in vitro on unlabeled hydrogels and hydrogels labeled at different concentrations, and in vivo on rats with unlabeled and labeled hydrogels implanted dorsally. In vivo MRI was performed at 1, 3, 5, and 7 weeks postimplantation. Implants were easily identified on T1-weighted SE, and fluid accumulation from inflammation was distinguished on T2-weighted turbo SE. Implants were segmented on contiguous T1-weighted SPGR slices using a threshold of 1.8 times the background muscle signal intensity; implant volume and mean T1 values were then calculated at each timepoint. Histopathology was performed on implants in the same plane as MRI and compared to imaging results. STATISTICAL TESTS: Unpaired t-tests and one-way analysis of variance (ANOVA) were used for comparisons. A P value <0.05 was considered to be statistically significant. RESULTS: Hydrogel labeling with MnP resulted in a significant T1 reduction in vitro (T1 = 517 ± 36 msec vs. 879 ± 147 msec unlabeled). Mean T1 values of labeled implants in rats increased significantly by 23% over time, from 1 to 7 weeks postimplantation (651 ± 49 msec to 801 ± 72 msec), indicating decreasing implant density. DATA CONCLUSION: Polymer-binding MnP enables in vivo tracking of vinyl-group coupling polymers. EVIDENCE LEVEL: 1. TECHNICAL EFFICACY: Stage 1.

SLERT, as a novel biomarker, orchestrates endometrial cancer metastasis via regulation of BDNF/TRKB signaling.

BACKGROUND: Recent evidence suggests that the box H/ACA small nucleolar RNA (snoRNA)-ended long noncoding RNA (lncRNA), SLERT, plays a critical role in gene regulation. However, its role in cancer remains undetermined. Herein, we explored its implication in human endometrial cancer (EC). METHODS: EC plasma and tissue samples were collected for the detection of SLERT expression using qRT-PCR method. The functional investigation was tested by CCK-8 and transwell assays. Luciferase reporter, RNA pull-down, and immunoprecipitation (RIP) assays were used to determine the regulatory network involved in SLERT. The in vivo effect of SLERT was tested by caudal vein lung metastasis model. RESULTS: Stable knockdown of SLERT significantly inhibited EC cell (KLE and AN3CA) migration and invasion, while it did not affect cell viability. SLERT induced epithelial-mesenchymal transition (EMT) via elevating N-cadherin and Vimentin and downregulating E-cadherin. Further investigation showed that SLERT directly binds to METTL3, increasing the m6A levels of BDNF mRNA; then, the m6A sites were read by IGF2BP1, enhancing BDNF mRNA stability, followed by the activation of BDNF/TRKB signaling, an inducer of EMT. The animal model showed that overexpression of SLERT increased EC cell lung metastasis, and this effect was effectively blocked by BDNF silencing or treatment with TRKB inhibitor k252a. Clinically, EC patients have high levels of SLERT both in tissue or plasma, which might be used as a biomarker of diagnosis and prognosis. CONCLUSION: Our findings, for the first time, uncover the metastasis-promoting effect of SLERT in EC via in vitro and in vivo evidence, providing a potential therapeutic target for metastatic EC treatment.

A Manganese Porphyrin Platform for the Design and Synthesis of Molecular and Targeted MRI Contrast Agents.

Magnetic resonance imaging (MRI) contrast agents, in contrast to the plethora of fluorescent agents available to target disease biomarkers or exogenous implants, have remained predominantly non-specific. That is, they do not preferentially accumulate in specific locations in vivo because doing so necessitates longer contrast retention, which is contraindicated for current gadolinium (Gd) agents. This double-edge sword implies that Gd agents can offer either rapid elimination (but lack specificity) or targeted accumulation (but with toxicity risks). For this reason, MRI contrast agent innovation has been severely constrained. Gd-free alternatives based on manganese (Mn) chelates have been largely ineffective, as they are inherently unstable. In this study, we present a Mn(III) porphyrin (MnP) platform for bioconjugation, offering the highest stability and chemical versatility compared to any other T1 contrast agent. We exploit the inherent metal stability conferred by porphyrins and the absence of pendant bases (found in Gd or Mn chelates) that limit versatile functionalization. As proof-of-principle, we demonstrate labeling of human serum albumin, a model protein, and collagen hydrogels for applications in in-vivo targeted imaging and material tracking, respectively. In-vitro and in-vivo results confirm unprecedented metal stability, ease of functionalization, and high T1 relaxivity. This new platform opens the door to ex-vivo validation by fluorescent imaging and multipurpose molecular imaging in vivo.

Primer and Historical Review on Rapid Cardiac CINE MRI.

Acceleration is an important consideration when imaging moving organs such as the heart. Not only does acceleration enable motion-free scans but, more importantly, it lies at the heart of capturing the dynamics of cardiac motion. For over three decades, various ingenious approaches have been devised and implemented for rapid CINE MRI suitable for dynamic cardiac imaging. Virtually all techniques relied on acquiring less data to reduce acquisition times. Parallel imaging was among the first of these innovations, using multiple receiver coils and mathematical algorithms for reconstruction; acceleration factors of 2 to 3 were readily achieved in clinical practice. However, in the context of imaging dynamic events, further decreases in scan time beyond those provided by parallel imaging were possible by exploiting temporal coherencies. This recognition ushered in the era of k-t accelerated MRI, which utilized predominantly statistical methods for image reconstruction from highly undersampled k-space. Despite the successes of k-t acceleration methods, however, the accuracy of reconstruction was not always guaranteed. To address this gap, MR physicists and mathematicians applied compressed sensing theory to ensure reconstruction accuracy. Reconstruction was, indeed, more robust, but it required optimizing regularization parameters and long reconstruction times. To solve the limitations of all previous methods, researchers have turned to artificial intelligence and deep neural networks for the better part of the past decade, with recent results showing rapid, robust reconstruction. This review provides a comprehensive overview of key developments in the history of CINE MRI acceleration, and offers a unique and intuitive explanation behind the techniques and underlying mathematics.Level of Evidence: 5Technical Efficacy Stage: 1.

Three-Dimensional Bioprinted MR-Trackable Regenerative Scaffold for Postimplantation Monitoring on T1-Weighted MRI.

BACKGROUND: A three-dimensional (3D) bioprinted tissue scaffold is a promising therapeutic that goes beyond providing physical support for tissue regeneration by enabling precise spatial control over scaffold geometry and integration of different materials/cells. Critically important is in vivo confirmation of correct scaffold placement and retention during the initial 24 hours postimplantation, to detect unwanted implant migration. PURPOSE: To incorporate a safe, efficient MR contrast agent into a bioprinting workflow, and to achieve bright-contrast scaffold monitoring in vivo postimplantation. STUDY TYPE: In vitro and animal in vivo longitudinal study. ANIMAL MODEL: Two female Sprague Dawley rats (~200 g) for labeled and unlabeled scaffold implantation in the subcutaneous dorsal space flanking the vertebral column. FIELD STRENGTH/SEQUENCE: A 7.0 T/T1 -weighted spin echo (SE) sequence and T1 mapping using turbo SE with variable repetition times (TRs). ASSESSMENT: Cell viability and proliferation were assessed over 2 weeks after labeling bioprinted gelatin/alginate scaffolds with MnPNH2 (0.5 mM, 24 hours). In vitro MRI was performed 0, 12, and 24 hours postlabeling in nine labeled and three unlabeled (control) scaffolds to monitor T1 evolution. In vivo MRI was performed immediately and 24 hours postimplantation to assess T1 . Acute inflammation near surgical site was monitored in one rat to 3 days. STATISTICAL TESTS: One-way analysis of variance with Tukey-Kramer post hoc analysis (P < 0.01). RESULTS: Cell viability was unaffected by bioprinting/labeling: viability exceeded 90% in all scaffolds after 1 week. In vitro T1 's were significantly lower in labeled scaffolds compared to control (207 msec vs. 2257 msec) immediately postlabeling and 24 hours later (1227 msec vs. 2257 msec). In vivo T1 's were significantly different (243.6 msec vs. 2414.6 msec) immediately postimplantation, and no differences emerged compared to respective in vitro control/labeled counterparts. The 24-hours imaging and gross pathology confirmed migration of scaffolds beyond the imaging field. DATA CONCLUSION: We report an MR-detectable, cell-compatible bioprinted scaffold, utilizing a T1 -weighting contrast agent for high-resolution, postimplantation scaffold tracking. EVIDENCE LEVEL: 2 TECHNICAL EFFICACY: Stage 1.

MRI method for labeling and imaging decellularized extracellular matrix scaffolds for tissue engineering.

PURPOSE: To develop a facile method for labeling and imaging decellularized extracellular matrix (dECM) scaffolds intended for regenerating 3D tissues. METHODS: A small molecule manganese porphyrin, MnPNH2 , was synthesized and used to label dECM scaffolds made from porcine bladder and trachea and murine whole lungs. The labeling protocol was optimized on bladder dECM, and imaging on a 3T clinical scanner was performed to assess reductions in T1 and T2 relaxation times. In vivo MRI was performed on dECM injected in the rat dorsum to verify sensitivity of detection. Toxicity assays for cell viability, metabolism, and proliferation were performed on human umbilical vein endothelial cells. The incorporation of MnPNH2 and its long-term retention in dECM were assessed on transmission electron microscopy and ultraviolet absorbance of eluted MnPNH2 over time. RESULTS: All tissues, including thick whole 3D organs, were uniformly labeled and demonstrated high signal-to-noise on MRI. A nearly 10-fold reduction in T1 was consistently obtained at a labeling dose of 0.4 mM, and even 0.2 mM provided sufficient contrast in vivo and ex vivo. No toxicity was observed up to 0.4 mM, the maximum tested. Binding studies suggested nonspecific association, and retention studies in the labeled whole decellularized lungs revealed less than 20% MnPNH2 loss over 30 days, the majority occurring in the first 3 days after labeling. CONCLUSION: The proposed labeling method is the first report for visualizing dECM on MRI and has the potential for long-term monitoring and optimization of dECM-based organ tissue engineering.

Heart failure with preserved ejection fraction: the missing pieces in diagnostic imaging.

Heart failure with preserved ejection fraction (HFpEF) is an increasingly prevalent phenotype affecting over half of today's heart failure patients. With no proven therapy and no universally accepted diagnostic guideline, many HFpEF patients continue to be misdiagnosed or underdiagnosed at the early stages until the disease has progressed much further along. It is extremely difficult to diagnose the HFpEF patient, because they have a normal ejection fraction and present with non-specific symptoms such as dyspnea or exercise intolerance. To provide greater specificity, the current diagnostic criteria mandate the presence of diastolic dysfunction, where myocardial relaxation is impaired and ventricular filling pressure is elevated as a result of a hypertrophic and stiff heart. Unfortunately, diastolic dysfunction reflects late-stage structural and functional changes and offers a very narrow window, if at all, for successful intervention. In this article, we review the imaging modalities used in the current diagnostic workflow for assessing HFpEF. We also describe the most up-to-date insight into its pathophysiological basis, which attributes systemic inflammation driven by comorbidities as the initiator of disease. With this extramyocardial perspective, we provide our recommendation on new imaging targets that extend beyond the heart to enable early, accurate diagnosis of HFpEF and allow an opportunity for treating this fatal condition.

miR-125a-5p post-transcriptionally suppresses GALNT7 to inhibit proliferation and invasion in cervical cancer cells via the EGFR/PI3K/AKT pathway.

BACKGROUND: The carcinogenesis and progression of cervical cancer is a complex process in which numerous microRNAs are involved. The purpose of this study is to investigate the role of miR-125a-5p in progression of cervical cancer. METHODS: RT-qPCR was used to detect the expression of miR-125a-5p and GALNT7 in cervical cancer tissues and cell lines. Then, the miR-125a-5p mimic, miR-125a-5p inhibitor, GALNT7 siRNA, or/and pcDNA-GALNT7 were respectively transfected into HeLa and Caski cervical cancer cells, and Cell Counting kit-8 assay, Transwell assay and flow cytometry analysis were respectively used to observe cell proliferation, invasion and apoptosis. Subsequently, luciferase reporter gene assay was employed in confirming the target relationship between miR-125a-5p and GALNT7. MiR-125a-5p mimic or/and pcDNA-GALNT7 were transfected into the cervical cancer cells at the absence of epidermal growth factor (EGF) or not, and the pcDNA-GALNT7 was transfected into the cervical cancer cells at the absence of inhibitors of multiple kinases or not. Furthermore, the effect of miR-125a-5p on tumor growth was also studied using a xenograft model of nude mice. RESULTS: MiR-125a-5p was down-regulated in both cervical cancer tissues and cell lines and it inhibited cell proliferation and invasion of cervical cancer cells. MiR-125a-5p directly targeted and post-transcriptionally downregulated GALNT7 that was strongly upregulated in cervical cancer tissues and cell lines. Similar to the effect of miR-125a-5p mimic, silencing GALNT7 inhibited proliferation and invasion of cervical cancer cells. In addition, miR-125a-5p overexpression could counteract both GALNT7- and EGF-induced cell proliferation and invasion. GALNT7 promoted cell proliferation and invasion by activating the EGFR/PI3K/AKT kinase pathway, which could be abated by the inhibitors of the kinases. Moreover, the role of miR-125a-5p inhibited tumor formation in cervical cancer by suppressing the expression of GALNT7 in vivo. CONCLUSION: In conclusion, miR-125a-5p suppressed cervical cancer progression by post-transcriptionally downregulating GALNT7 and inactivating the EGFR/PI3K/AKT pathway.

Roles of a TMPO-AS1/microRNA-200c/TMEFF2 ceRNA network in the malignant behaviors and 5-FU resistance of ovarian cancer cells.

Competing endogenous RNA (ceRNA) networks consisted of long non-coding RNA (lncRNA), microRNA (miRNA) and mRNAs have aroused great interests recently. The current study aims to probe the mechanisms of lncRNA TMPO-AS1 in ovarian cancer (OC) development. A 5-fluorouracil (5-FU)-resistant subline of OC SKOV3 cells was developed, and differentially expressed lncRNAs in OC tissues and SKOV3 cells were analyzed. The miRNAs, genes and signaling pathways interacted with TMPO-AS1 were predicted and validated. TMPO-AS1 and the validated miRNA were inhibited to analyze their roles in malignant behaviors and 5-FU resistance of OC cells. In vivo studies were performed by inducing xenograft tumors in nude mice. Consequently, TMPO-AS1 was highly expressed in OC tissues and SKOV3 cells. TMPO-AS1 regulated transmembrane protein with epidermal growth factor and two follistatin motifs 2 (TMEFF2) through sponging miR-200c in OC cells, during which the PI3K/Akt signaling pathway was activated. Silenced TMPO-AS1 and over-expressed miR-200c inhibited epithelial-mesenchymal transition (EMT), invasion, migration and 5-FU resistance of OC cells. This study demonstrated that silencing of TMPO-AS1 might attenuate OC progression through inhibiting the invasion, metastasis and drug resistance of OC cells via the miR-200c/TMEFF2 network and the disruption of the PI3K/Akt signaling pathway.

PI3K-p110α mediates the oncogenic activity induced by loss of the novel tumor suppressor PI3K-p85α.

Mutation or loss of the p85 regulatory subunit of phosphatidylinositol 3-kinase (PI3K) is emerging as a transforming factor in cancer, but the mechanism of transformation has been controversial. Here we find that hemizygous deletion of the PIK3R1 gene encoding p85α is a frequent event in breast cancer, with PIK3R1 expression significantly reduced in breast tumors. PIK3R1 knockdown transforms human mammary epithelial cells, and genetic ablation of Pik3r1 accelerates a mouse model of HER2/neu-driven breast cancer. We demonstrate that partial loss of p85α increases the amount of p110α-p85 heterodimers bound to active receptors, augmenting PI3K signaling and oncogenic transformation. Pan-PI3K and p110α-selective pharmacological inhibition effectively blocks transformation driven by partial p85α loss both in vitro and in vivo. Together, our data suggest that p85α plays a tumor-suppressive role in transformation, and suggest that p110α-selective therapeutics may be effective in the treatment of breast cancer patients with PIK3R1 loss.

The p110α and p110ß isoforms of PI3K play divergent roles in mammary gland development and tumorigenesis.

Class Ia phosphatidylinositol 3 kinase (PI3K) is required for oncogenic receptor-mediated transformation; however, the individual roles of the two commonly expressed class Ia PI3K isoforms in oncogenic receptor signaling have not been elucidated in vivo. Here, we show that genetic ablation of p110α blocks tumor formation in both polyoma middle T antigen (MT) and HER2/Neu transgenic models of breast cancer. Surprisingly, p110ß ablation results in both increased ductal branching and tumorigenesis. Biochemical analyses suggest a competition model in which the less active p110ß competes with the more active p110α for receptor binding sites, thereby modulating the level of PI3K activity associated with activated receptors. Our findings demonstrate a novel p110ß-based regulatory role in receptor-mediated PI3K activity and identify p110α as an important target for treatment of HER2-positive disease.

Human bronchial carcinoid tumor initiating cells are targeted by the combination of acetazolamide and sulforaphane.

BACKGROUND: Bronchial carcinoids are neuroendocrine tumors that present as typical (TC) and atypical (AC) variants, the latter being more aggressive, invasive and metastatic. Studies of tumor initiating cell (TIC) biology in bronchial carcinoids has been hindered by the lack of appropriate in-vitro and xenograft models representing the bronchial carcinoid phenotype and behavior. METHODS: Bronchial carcinoid cell lines (H727, TC and H720, AC) were cultured in serum-free growth factor supplemented medium to form 3D spheroids and serially passaged up to the 3rd generation permitting expansion of the TIC population as verified by expression of stemness markers, clonogenicity in-vitro and tumorigenicity in both subcutaneous and orthotopic (lung) models. Acetazolamide (AZ), sulforaphane (SFN) and the AZ + SFN combination were evaluated for targeting TIC in bronchial carcinoids. RESULTS: Data demonstrate that bronchial carcinoid cell line 3rd generation spheroid cells show increased drug resistance, clonogenicity, and tumorigenic potential compared with the parental cells, suggesting selection and expansion of a TIC fraction. Gene expression and immunolabeling studies demonstrated that the TIC expressed stemness factors Oct-4, Sox-2 and Nanog. In a lung orthotopic model bronchial carcinoid, cell line derived spheroids, and patient tumor derived 3rd generation spheroids when supported by a stroma, showed robust tumor formation. SFN and especially the AZ + SFN combination were effective in inhibiting tumor cell growth, spheroid formation and in reducing tumor formation in immunocompromised mice. CONCLUSIONS: Human bronchial carcinoid tumor cells serially passaged as spheroids contain a higher fraction of TIC exhibiting a stemness phenotype. This TIC population can be effectively targeted by the combination of AZ + SFN. Our work portends clinical relevance and supports the therapeutic use of the novel AZ+ SFN combination that may target the TIC population of bronchial carcinoids.

Assessment of microvascular dysfunction in acute limb ischemia-reperfusion injury.

BACKGROUND: Ischemia-reperfusion (I/R) injury involves damage to the microvessel structure (eg, increased permeability) and function (blunted vasomodulation). While microstructural damage can be detected with dynamic contrast-enhanced (DCE) MRI, there is no diagnostic to detect deficits in microvascular function. PURPOSE: To apply a novel MRI method for evaluating dynamic vasomodulation to assess microvascular dysfunction in skeletal muscle following I/R injury. STUDY TYPE: Prospective, longitudinal. ANIMAL MODEL: Twenty-three healthy male adult Sprague-Dawley rats. FIELD STRENGTH/SEQUENCE: Dynamic T1 fast field echo imaging at 3.0T with preinjection T1 mapping. ASSESSMENT: Injury in the left hindlimb was induced using a 3-hour I/R procedure. Longitudinal MRI scanning was performed up to 74 days, with animals completing assessment at different intervals for histological and laser Doppler perfusion validation. Pharmacokinetic parameters Ktrans and ve were determined following i.v. injection of gadovist (0.1 mmol/kg). Vasomodulatory response was probed on gadofosveset (0.3 mmol/kg) using hypercapnic gases delivered through a controlled gas-mixing circuit to induce vasoconstriction and vasodilation in ventilated rats. Heart rate and blood oxygen saturation were monitored. STATISTICAL TESTS: Two-way analysis of variance with Tukey-Kramer post-hoc analysis was used to determine significant changes in vasomodulatory response, Ktrans , and ve . RESULTS: This new MRI technique revealed impaired vasomodulation in the injured hindlimb. Vasoconstriction was maintained, but vasodilation was blunted up to 21 days postinjury (P < 0.05). However, DCE-MRI measured Ktrans and ve were significantly (P < 0.05) different from baseline only during acute inflammation (Day 3), with severe inflammation noted on histology. DATA CONCLUSION: While conventional DCE-MRI shows normalization after the acute phase, our new approach reveals sustained functional impairment in muscle microvasculature following I/R injury, with compromised response in vasomotor tone present for at least 21 days. LEVEL OF EVIDENCE: 4 Technical Efficacy: Stage 1 J. Magn. Reson. Imaging 2019;49:1174-1185.

H19 knockdown suppresses proliferation and induces apoptosis by regulating miR-148b/WNT/ß-catenin in ox-LDL -stimulated vascular smooth muscle cells.

BACKGROUND: Long non-coding RNAs (lncRNAs) have been identified as critical regulators in the development of atherosclerosis (AS). Here, we focused on discussing roles and molecular mechanisms of lncRNA H19 in vascular smooth muscle cells (VSMCs) progression. METHODS: RT-qPCR assay was used to detect the expression patterns of H19 and miR-148b in clinical samples and cells. Cell proliferative ability was evaluated by CCK-8 and colony formation assays. Cell apoptotic capacity was assessed by apoptotic cell percentage and the caspase-3 activity. Bioinformatics analysis, luciferase and RNA immunoprecipitation (RIP) assays were employed to demonstrate cell percentage and the relationship among H19, miR-148b and wnt family member 1 (WNT1). Western blot assay was performed to determine expressions of proliferating cell nuclear antigen (PCNA), ki-67, Bax, Bcl-2, WNT1, ß-catenin, C-myc and E-cadherin. RESULTS: The level of H19 was increased and miR-148b expression was decreased in human AS patient serums and oxidized low-density lipoprotein (ox-LDL)-stimulated human aorta vascular smooth muscle cells (HA-VSMCs). H19 knockdown suppressed proliferation and promoted apoptosis in HA-VSMCs following the treatment of ox-LDL. H19 inhibited miR-148b expression by direct interaction. Moreover, miR-148b inhibitor could reverse the effects of H19 depletion on proliferation and apoptosis in ox-LDL-stimulated HA-VSMCs. Further mechanical explorations showed that WNT1 was a target of miR-148b and H19 acted as a competing endogenous RNA (ceRNA) of miR-148b to enhance WNT1 expression. Furthermore, miR-148 inhibitor exerted its pro-proliferation and anti-apoptosis effects through activating WNT/ß-catenin signaling in ox-LDL-stimulated HA-VSMCs. CONCLUSION: H19 facilitated proliferation and inhibited apoptosis through modulating WNT/ß-catenin signaling pathway via miR-148b in ox-LDL-stimulated HA-VSMCs, implicating the potential values of H19 in AS therapy.

Chemopreventive effects of aspirin at a glance.

Experimental, epidemiological, and clinical data from the last two decades have each supported the hypothesis that aspirin possesses anticancer properties, and that its use may also reduce the lifetime probability of developing or dying from a number of cancers. Aspirin's ability to act on multiple key metabolic and signaling pathways via inhibition of the cyclooxygenase (COX) enzyme, as well as through COX-independent mechanisms, makes it particularly relevant in the fight against cancer. A growing body of evidence indicates that aspirin may not only reduce cancer risk, but also prevent metastasis and angiogenesis while slowing the rate of mutation-inducing DNA damage. These emerging benefits of aspirin are offset to some extent by the known risks of treatment, such as cardiovascular events and gastrointestinal bleeding. However, it has been shown that pre-treatment risk assessment of individual patients and the use of proton pump inhibitors or Helicobacter pylori eradication therapy concomitantly with aspirin treatment can reduce these potential risks. Thus, the significant benefits of aspirin treatment, coupled with recent data concerning its risks, may prove to tip the balance in favor of aspirin use in cancer prevention.

Combined inhibition of PI3K and PARP is effective in the treatment of ovarian cancer cells with wild-type PIK3CA genes.

OBJECTIVE: Combined inhibition of PI3K and PARP has been shown to be effective in the treatment of preclinical models of breast cancer and prostate cancer independent of BRCA or PIK3CA mutational status. However, the knowledge about this combination treatment in ovarian cancer is limited. The aim of this study was to evaluate the therapeutic effect of PI3K inhibitor BKM120 and PARP inhibitor Olaparib on ovarian cancer cell lines bearing wild-type PIK3CA genes. METHODS: We exposed three wild-type PIK3CA ovarian cancer cell lines to a PI3K inhibitor BKM120 and/or a PARP inhibitor Olaparib. The effect of BKM120 as a single-agent or in combination with Olaparib was evaluated by Cell Count Kit (CCK8) assay, immunoblotting, comet assay, flow cytometry and immunofluorescence staining assay. The combination indexes for synergistic effect on cell viability were calculated with the Chou-Talalay method. Ex vivo cultured ovarian cancer tissues from patients were analyzed by histological and immunohistochemical analyses. RESULTS: Combined inhibition of PI3K and PARP effectively synergized to block the growth of three wild-type PIK3CA ovarian cancer cell lines and explants of a primary ovarian tumor specimen. Mechanistically, dual blockade of PI3K and PARP in these ovarian cancer cell lines resulted in substantially attenuated PI3K/AKT/mTOR signaling, impaired DNA damage response and deficient homologous recombination repair, with remarkable BRCA downregulation. CONCLUSIONS: The combined use of PI3K inhibitor BKM120 and PARP inhibitor Olaparib may be effective in ovarian cancers with a broader spectrum of cancer-associated genetic alterations but not limited to those with mutant PIK3CA or BRCA genes. BRCA downregulation may be a potential biomarker for the effective response to the proposed combination treatment.

Mutant PIK3CA accelerates HER2-driven transgenic mammary tumors and induces resistance to combinations of anti-HER2 therapies.

Human epidermal growth factor receptor 2 (HER2; ERBB2) amplification and phosphatidylinositol-4,5-bisphosphate 3-kinase, catalytic subunit alpha (PIK3CA) mutations often co-occur in breast cancer. Aberrant activation of the phosphatidylinositol 3-kinase (PI3K) pathway has been shown to correlate with a diminished response to HER2-directed therapies. We generated a mouse model of HER2-overexpressing (HER2(+)), PIK3CA(H1047R)-mutant breast cancer. Mice expressing both human HER2 and mutant PIK3CA in the mammary epithelium developed tumors with shorter latencies compared with mice expressing either oncogene alone. HER2 and mutant PIK3CA also cooperated to promote lung metastases. By microarray analysis, HER2-driven tumors clustered with luminal breast cancers, whereas mutant PIK3CA tumors were associated with claudin-low breast cancers. PIK3CA and HER2(+)/PIK3CA tumors expressed elevated transcripts encoding markers of epithelial-to-mesenchymal transition and stem cells. Cells from HER2(+)/PIK3CA tumors more efficiently formed mammospheres and lung metastases. Finally, HER2(+)/PIK3CA tumors were resistant to trastuzumab alone and in combination with lapatinib or pertuzumab. Both drug resistance and enhanced mammosphere formation were reversed by treatment with a PI3K inhibitor. In sum, PIK3CA(H1047R) accelerates HER2-mediated breast epithelial transformation and metastatic progression, alters the intrinsic phenotype of HER2-overexpressing cancers, and generates resistance to approved combinations of anti-HER2 therapies.

Role of SDF-1 and CXCR4 in the proliferation, migration and invasion of cervical cancer.

This study was to investigate the role of stromal cell-derived factor 1 (SDF-1) and its corresponding receptor CXCR4 in the proliferation, migration and invasion of cervical cancer HeLa cells. CXCR4 expression in HeLa cells was measured by flow cytometry and Western Blot. Role of SDF-1 and CXCR4 in the HeLa cells proliferation was measured by MTT. Role of SDF-1 and CXCR4 in the migration and invasion of HeLa cell was measured by Boyden chamber. High expression of CXCR4 was observed on the surface of HeLa cells. Proliferation ability of HeLa cells was significantly increased after SDF-1 stimulation, which showed dose-dependent manner. After knock-down of CXCR4 expression by RNAi, SDF-1-stimulated HeLa cells proliferation was significantly blocked (P<0.05). SDF-1 can induce migration and invasion of Hela cells, SDF-1-stimulated HeLa cells migration and invasion was significantly blocked (P<0.05) after knock-down of CXCR4 expression by RNAi. High expression of surface CXCR4 plays an important role in the proliferation, migration and invasion of HeLa cells.