<div>AbstractPurpose:<p>Clear cell renal cell carcinoma (ccRCC) is frequently associated with inactivation of the von Hippel–Lindau tumor suppressor, resulting in activation of HIF-1α and HIF-2α. The current paradigm, established using mechanistic cell-based studies, supports a tumor promoting role for HIF-2α, and a tumor suppressor role for HIF-1α. However, few studies have comprehensively examined the clinical relevance of this paradigm. Furthermore, the hypoxia-associated factor (HAF), which regulates the HIFs, has not been comprehensively evaluated in ccRCC.</p>Experimental Design:<p>To assess the involvement of HAF/HIFs in ccRCC, we analyzed their relationship to tumor grade/stage/outcome using tissue from 380 patients, and validated these associations using tissue from 72 additional patients and a further 57 patients treated with antiangiogenic therapy for associations with response. Further characterization was performed using single-cell mRNA sequencing (scRNA-seq), RNA-<i>in situ</i> hybridization (RNA-ISH), and IHC.</p>Results:<p>HIF-1α was primarily expressed in tumor-associated macrophages (TAMs), whereas HIF-2α and HAF were expressed primarily in tumor cells. TAM-associated HIF-1α was significantly associated with high tumor grade and increased metastasis and was independently associated with decreased overall survival. Furthermore, elevated TAM HIF-1α was significantly associated with resistance to antiangiogenic therapy. In contrast, high HAF or HIF-2α were associated with low grade, decreased metastasis, and increased overall survival. scRNA-seq, RNA-ISH, and Western blotting confirmed the expression of HIF-1α in M2-polarized CD163-expressing TAMs.</p>Conclusions:<p>These findings highlight a potential role of TAM HIF-1α in ccRCC progression and support the reevaluation of HIF-1α as a therapeutic target and marker of disease progression.</p></div>

El objetivo de este artículo fue diseñar una secuencia didáctica para el tratamiento del tema de los circuitos eléctricos con una metodología de aprendizaje activo para dar respuesta a la pregunta sobre qué tipo de aprendizaje se promueve en los estudiantes utilizando esas metodologías en las aulas virtuales de las clases de física en ingeniería y su posterior aplicación en modalidad híbrida. El estudio es cualitativo y sigue las fases de la investigación-acción; para la construcción del problema contextualizado, se usó una metodología DIPCING, diseñada ex profeso para la enseñanza de la ingeniería, y se propuso la infografía como producto de aprendizaje integrador. Se empleó un instrumento denominado “aplicación a la ingeniería” para cuantificar habilidades de orden superior. Con el diseño y la evaluación que se documenta, se aportan herramientas para la implementación del ABP en carreras de ingeniería en una modalidad híbrida. Con esta propuesta, los alumnos comprenden conocimientos científicos (sobre circuitos eléctricos) y tienen nociones de cómo utilizarlos en el entorno laboral.

<div>Abstract<p>Cancer cells respond to hypoxia by upregulating the hypoxia-inducible factor 1α (HIF1A) transcription factor, which drives survival mechanisms that include metabolic adaptation and induction of angiogenesis by VEGF. Pancreatic tumors are poorly vascularized and severely hypoxic. To study the angiogenic role of HIF1A, and specifically probe whether tumors are able to use alternative pathways in its absence, we created a xenograft mouse tumor model of pancreatic cancer lacking HIF1A. After an initial delay of about 30 days, the HIF1A-deficient tumors grew as rapidly as the wild-type tumors and had similar vascularization. These changes were maintained in subsequent passages of tumor xenografts <i>in vivo</i> and in cell lines <i>ex vivo</i>. There were many cancer cells with a "clear-cell" phenotype in the HIF1A-deficient tumors; this was the result of accumulation of glycogen. Single-cell RNA sequencing (scRNA-seq) of the tumors identified hypoxic cancer cells with inhibited glycogen breakdown, which promoted glycogen accumulation and the secretion of inflammatory cytokines, including interleukins 1β (IL1B) and 8 (IL8). scRNA-seq of the mouse tumor stroma showed enrichment of two subsets of myeloid dendritic cells (cDC), cDC1 and cDC2, that secreted proangiogenic cytokines. These results suggest that glycogen accumulation associated with a clear-cell phenotype in hypoxic cancer cells lacking HIF1A can initiate an alternate pathway of cytokine and DC-driven angiogenesis. Inhibiting glycogen accumulation may provide a treatment for cancers with the clear-cell phenotype.</p>Significance:<p>These findings establish a novel mechanism by which tumors support angiogenesis in an HIF1α-independent manner.</p></div>

<p>Supplementary Methods 1. Tumor Dissociation and Single Cell RNA Sequencing. Supplementary Figure S1. Sequences of shRNAs used to inhibit tumor HIF1A and GYS1. Supplementary Figure S2. Validation of HIF1A knockdown and inhibition of HIF1A target gene expression. Supplementary Figure S3. Pimonidazole staining for hypoxia and Oil Red O for lipid in shHIF1A tumors. Supplementary Figure S4. Characteristics of shGYS1 and shHIF1A cells and tumors. Supplementary Figure S5. IL-1B expression is maintained in second generation growth of shHIF1A xenografts. Supplementary Table 1. shHIF1A IL1B and NF-κB activation signatures Supplementary Table S2. Flow cytometry characterization of TIL in EV and shHIF1A tumors. Supplementary Table S3. Transcripts used to characterize mouse scRNAseq clusters cDC1 and cDC2.</p>

<p>Supplementary Methods 1. Tumor Dissociation and Single Cell RNA Sequencing. Supplementary Figure S1. Sequences of shRNAs used to inhibit tumor HIF1A and GYS1. Supplementary Figure S2. Validation of HIF1A knockdown and inhibition of HIF1A target gene expression. Supplementary Figure S3. Pimonidazole staining for hypoxia and Oil Red O for lipid in shHIF1A tumors. Supplementary Figure S4. Characteristics of shGYS1 and shHIF1A cells and tumors. Supplementary Figure S5. IL-1B expression is maintained in second generation growth of shHIF1A xenografts. Supplementary Table 1. shHIF1A IL1B and NF-κB activation signatures Supplementary Table S2. Flow cytometry characterization of TIL in EV and shHIF1A tumors. Supplementary Table S3. Transcripts used to characterize mouse scRNAseq clusters cDC1 and cDC2.</p>