Intraoperative identification of carcinoma at lumpectomy margins would enable reduced re-excision rates, which are currently as high as 20% to 50%. Although imaging of disease-associated biomarkers ...can identify malignancies with high specificity, multiplexed imaging of such biomarkers is necessary to detect molecularly heterogeneous carcinomas with high sensitivity. We have developed a Raman-encoded molecular imaging (REMI) technique in which targeted nanoparticles are topically applied on excised tissues to enable rapid visualization of a multiplexed panel of cell surface biomarkers at surgical margin surfaces. A first-ever clinical study was performed in which 57 fresh specimens were imaged with REMI to simultaneously quantify the expression of four biomarkers HER2, ER, EGFR, and CD44. Combined detection of these biomarkers enabled REMI to achieve 89.3% sensitivity and 92.1% specificity for the detection of breast carcinoma. These results highlight the sensitivity and specificity of REMI to detect biomarkers in freshly resected tissue, which has the potential to reduce the rate of re-excision procedures in cancer patients.
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HPV-16E7 is a major transforming protein, which has been implicated in the development of cervical cancer. The stability of E7 is thus important to ensure its fully functional status. Using the yeast ...two-hybrid system, we found that USP11 (ubiquitin-specific protease 11), a member of a protein family that cleaves polyubiquitin chains and/or ubiquitin precursors, interacts and forms a specific complex with HPV-16E7. Our results indicate that the USP11 can greatly increase the steady state level of HPV-16E7 by reducing ubiquitination and attenuating E7 degradation. In contrast, a catalytically inactive mutant of USP11 abolished the deubiquitinating ability and returned E7 to a normal rate of degradation. Moreover, USP11 not only protected E7 from ubiquitination but also influenced E7 function as a modulator of cell growth status. These results suggest that USP11 plays an important role in regulating the levels of E7 protein and subsequently affects the biological function of E7 as well as its contribution to cell transformation by HPV-16E7.
Surface-enhanced Raman scattering (SERS) nanoparticles (NPs) are increasingly being engineered for a variety of disease-detection and treatment applications. For example, we have previously developed ...a fiber-optic Raman-encoded molecular imaging (REMI) system for spectral imaging of biomarker-targeted SERS NPs topically applied on tissue surfaces to identify residual tumors at surgical margins. Although accurate tumor detection was achieved, the commercial SERS NPs used in our previous studies lacked the signal strength to enable high-speed imaging with high pixel counts (large fields of view and/or high spatial resolution), which limits their use for certain time-constrained clinical applications. As a solution, we explored the use of surface-enhanced resonant Raman scattering (SERRS) NPs to enhance imaging speeds. The SERRS NPs were synthesized de novo, and then conjugated to HER2 antibodies to achieve high binding affinity, as validated by flow cytometry. Under identical tissue-staining and imaging conditions, the targeted SERRS NPs enabled reliable identification of HER2-overexpressed tumor xenografts with 50-fold-enhanced imaging speed compared with our standard targeted SERS NPs. This enables our REMI system to image tissue surfaces at a rate of 150 cm2 per minute at a spatial resolution of 0.5 mm.
Krüppel-like factor 10 (KLF10) is a phospho-regulated transcriptional factor involved in many biological processes including lipogenesis; however, the transcriptional regulation on lipogenesis by ...KLF10 remains largely unclear. Lipogenesis is important in the development of nonalcoholic fatty liver disease (NAFLD) which was known regulated mainly by AMP-activated protein kinase (AMPK) and sterol regulatory element-binding protein (SREBP-1C). Interesting, our previous study using phosphorylated site prediction suggested a regulation of AMPK on KLF10. Therefore, we aimed to study the protein–protein interactions of AMPK on the regulation of KLF10, and to delineate the mechanisms of phosphorylated KLF10 in the regulation of NAFLD through SREBP-1C. We performed
in vitro
and
in vivo
assays that identified AMPK phosphorylates KLF10 at Thr189 and subsequently modulates the steady state level of KLF10. Meanwhile, a chromatin immunoprecipitation–chip assay revealed the novel target genes and signaling cascades of corresponding to phosphorylated KLF10. SREBP-1C was identified as a target gene suppressed by phosphorylated KLF10 through promoter binding. We further performed high-fat-diet-induced NAFLD models using hepatic-specific KLF10 knockout mice and wild-type mice and revealed that KLF10 knockout markedly led to more severe NAFLD than that in wild-type mice. Taken together, our findings revealed for the first time that AMPK activates and stabilizes the KLF10 protein
via
phosphorylation at Thr189, thereby repressing the expression of SREBP-1C and subsequent lipogenesis pathways along with metabolic disorders. We suggested that the targeted manipulation of liver metabolism, particularly through increased KLF10 expression, is a potential alternative solution for treating NAFLD.
Background: Competition for freshwater between cities and agriculture is projected to grow due to rapid urbanization, particularly in the Global South. Water reallocation from rural to urban regions ...has become a common strategy to meet freshwater needs in growing cities. Conceptual issues and associated measurement problems have impeded efforts to compare and learn from global experiences. This review examines the status and trends of water reallocation from rural to urban regions based on academic literature and policy documents. Methods: We conduct a systematic literature review to establish the global reallocation database (GRaD). This process yielded 97 published studies (academic and policy) on rural-to-urban reallocation. We introduce the concept of reallocation 'dyads' as the unit of analysis to describe the pair of a recipient (urban) and donor (rural) region. A coding framework was developed iteratively to classify the drivers, processes and outcomes of water reallocation from a political economy perspective. Results: The systematic review identified 69 urban agglomerations receiving water through 103 reallocation projects (dyads). Together these reallocation dyads involve approximately 16 billion m3 of water per year moving almost 13 000 kilometres to urban recipient regions with an estimated 2015 population of 383 million. Documented water reallocation dyads are concentrated in North America and Asia with the latter constituting the majority of dyads implemented since 2000. Synthesis: The analysis illustrates how supply and demand interact to drive water reallocation projects, which can take many forms, although collective negotiation and administrative decisions are most prevalent. Yet it also reveals potential biases and gaps in coverage for parts of the Global South (particularly in South America and Africa), where reallocation (a) can involve informal processes that are difficult to track and (b) receives limited coverage by the English-language literature covered by the review. Data regarding the impacts on the donor region and compensation are also limited, constraining evidence to assess whether a water reallocation project is truly effective, equitable and sustainable. We identify frameworks and metrics for assessing reallocation projects and navigating the associated trade-offs by drawing on the concept of benefit sharing.
KLF10 is now classified as a member of the Krüppel-like transcription factor family and acts as a tumor suppressor. Although KLF10 is originally named as TGF-β-inducible early gene-1 and mimicking ...the anti-proliferative effect of TGF-β in various carcinoma cells, the transcriptional upregulatory function of KLF10 has been described for a variety of cytokines and in many diseases. Through in vivo and in vitro phosphorylation assays, we identified that KLF10 is a phosphorylated protein in cells. Using yeast-two hybrid screening and site direct mutagenesis, we also identified PIN1 as a novel KLF10 associated protein. PIN1 is a peptidyl-prolyl isomerase enzyme belonging to the parvulin family, which specifically recognizes phosphorylated Ser/Thr-Pro containing substrates. Through protein–protein interaction assays, we showed that the Pro-directed Ser/Thr-Pro motif at Thr-93 in the KLF10 N-terminal region is essential for the interaction between KLF10 and PIN1. More importantly, PIN1 interacts with KLF10 in a phosphorylation-dependent manner and this interaction promotes KLF10 protein degradation in cells. Therefore, KLF10 shows shorter protein stability compared with mutant KLF10 that lacks PIN1 binding ability after cycloheximide treatments. The reversely correlated expression profile between KLF10 and PIN1 as observed in cell lines was also shown in clinic pancreatic cancer specimen. Using in vitro kinase assays and depletion assays, we were able to show that RAF-1 phosphorylates the Thr-93 of KLF10 and affects the KLF10 expression level in cells. Thus these findings as a whole indicate that RAF-1 phosphorylation and PIN1 isomerization together regulate KLF10 stability and further affect the role of KLF10 in tumor progression.
•Human KLF10, a tumor suppressor, is one of the substrates of RAF1 kinase.•The Thr93 phosphorylation of KLF10 by RAF1 generates a PIN1-binding site.•The PIN1 binding increases the degradation of KLF10 in a phosphorylated manner.
Deregulation of transforming growth factor (TGF)-β function is a common feature of pancreatic cancer, rendering these cancers unresponsive to TGF-β–stimulated growth inhibition. Recent findings have ...supported a primary role for Krüppel-like factor 10 (KLF10) as an important transcription factor involved in mediating TGF-β1 signaling. The aim of this study was to evaluate the correlation between KLF10 expression and the clinical and pathologic features of pancreatic cancer. Tissue specimens from patients with pancreatic adenocarcinoma were retrospectively collected for immunohistochemical analysis. To demonstrate that Klf10 expression was primarily regulated by methylation status, the Klf10 promoter was examined by methylation-specific PCR using a pancreatic cancer cell line (Panc-1). DNA methyltransferase (DNMT) inhibitor and small-interfering RNA depletion of DNMT genes were used to reverse KLF10 expression in the Panc-1 cells. In parallel, DNMT1 expression was evaluated in the pancreatic cancer tissue specimens. In 95 pancreatic cancer tissue specimens, KLF10 expression was inversely correlated with pancreatic cancer stage ( P = 0.01). Multivariable analysis revealed that, in addition to the presence of distant metastasis at diagnosis ( P = 0.001 and 0.001, respectively), KLF10 was another independent prognostic factor related to progression-free and overall survival ( P = 0.018 and 0.037, respectively). The loss of KLF10 expression in advanced pancreatic cancer is correlated with altered methylation status, which seems to be regulated by DNMT1. Our results suggest that KLF10 is a potential clinical predictor for progression of pancreatic cancer.
Previous studies have shown that functionalized nanoparticles (NPs) topically applied on fresh tissues are able to rapidly target cell‐surface protein biomarkers of cancer. Furthermore, studies have ...shown that a paired‐agent approach, in which an untargeted NP is co‐administered with a panel of targeted NPs, controls for the nonspecific behavior of the NPs, enabling quantitative imaging of biomarker expression. However, given the complexities in nonspecific accumulation, diffusion, and chemical binding of targeted NPs in tissues, studies are needed to better understand these processes at the microscopic scale. Here, fresh tissues were stained with a paired‐agent approach, frozen, and sectioned to image the depth‐dependent accumulation of targeted and untargeted NPs. The ratio of targeted‐to‐untargeted NP concentrations—a parameter used to distinguish between tumor and benign tissues—was found to diminish with increasing NP diffusion depths due to nonspecific accumulation and poor washout. It was then hypothesized and experimentally demonstrated that larger NPs would exhibit less diffusion below tissue surfaces, enabling higher targeted‐to‐untargeted NP ratios. In summary, these methods and investigations have enabled the design of NP agents with improved sensitivity and contrast for rapid molecular imaging of fresh tissues.
A microscopic investigation of a nanoparticle‐based molecular imaging strategy is presented, which is used to inform the design of larger nanoparticles that exhibit reduced diffusion and improved binding to cell‐surface biomarkers when topically applied onto the surfaces of fresh tissue specimens. Data and preliminary modeling suggest the presence of a binding‐site barrier that restricts the diffusion of biomarker‐targeted nanoparticles in comparison to untargeted control nanoparticles.
► Studies on vascular development have revealed the crucial role TGF-β plays showing that TGF-β antibodies directly blocked vasculogenesis and angiogenesis. ► Although numerous studies have been ...done, the role of TGF-β as an angiogenic factor is still unclear. ► Our results provide first evidences to suggest that KLF10, a TGF-β-inducible early gene, regulates COX-1 transcripts expression and further affects tube formation in endothelial cells. ► Moreover, the KLF10 deficient mice exhibited decreased light transmission which represents the extend of platelet aggregation slowing down.
Krüppel-like family is a group of zinc-finger transcription factors which play key regulatory roles in cellular growth, development, differentiation and vascularization. Recent studies have shown that one of the members, KLF10, is specifically involved in the process of angiogenesis by acting as a key transcriptional regulator of TGF-β1 in pro-angiogenic cells differentiation and function. KLF10−/− mice also displayed impaired blood flow recovery after hindlimb ischemia. However, the mechanism of KLF10 induced angiogenesis is still not well understood. From ChIP-chip, which have been adopt to elucidate the novel target genes and signaling cascades of KLF10, COX-1 (also named as PTGS1) is one of the target genes that may be regulated by Klf10 through promoter binding. In order to investigate the function of KLF10/COX-1 axis, promoter activity, EMSA, ChIP-PCR and tube formation assays were serially performed. Our results demonstrated that KLF10 acts as a transcriptional activator on COX-1 promoter where overexpression of KLF10 induces COX-1 protein expression and mRNA expression in endothelial cells. It has been known that COX-1 is the key enzyme in prostaglandin biosynthesis which regulated angiogenesis in endothelial cells. Using tube formation assay, we further demonstrated that KLF10 overexpressed endothelial cells formed better organized three-dimensional tube structure in contrast to the control group did. To specifically investigate the role for KLF10 in angiogenesis, the its deficient mice exhibited decreased light transmission which represents the extend of platelet aggregation slowing down. Taken together, our results indicate an important role for KLF10 in angiogenesis through the activation of COX-1.
Sustainable and resilient food systems depend on sustainable and resilient water management. Resilience is characterised by overlapping decision spaces and scales and interdependencies among water ...users and competing sectors. Increasing water scarcity, due to climate change and other environmental and societal changes, makes putting caps on the consumption of water resources indispensable. Implementation requires an understanding of different domains, actors, and their objectives, and drivers and barriers to transformational change. We suggest a scale-specific approach, in which agricultural water use is embedded in a larger systems approach (including natural and human systems). This approach is the basis for policy coherence and the design of effective incentive schemes to change agricultural water use behaviour and, therefore, optimise the water we eat.
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