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Due to the limited efficacy and evident side effects of traditional chemotherapy drugs attributed to their lack of specificity and selectivity, novel strategies are essential for improving cancer treatment outcomes. Here, we successfully engineered Fe3O4magnetic nanoparticles coated with zeolitic imidazolate framework-8 (ZIF-8). The resulting nanocomposite (Fe3O4@ZIF-8) demonstrates efficient adsorption of a substantial amount of doxorubicin (DOX) due to the porous nature of ZIF-8. The drug-loaded nanoparticles, Fe3O4@ZIF-8/DOX, exhibit significant accumulation at the tumor site in SW620 colon-cancer-bearing mice when guided by an external magnetic field. Within the acidic microenvironment of the tumor, the ZIF-8 framework collapses, releasing DOX and effectively inducing tumor cell death, thereby inhibiting cancer progression while not causing undesired side effects, as confirmed by a variety of in vitro and in vivo characterizations. In comparison to free DOX, Fe3O4@ZIF-8/DOX nanoparticles show superior efficacy in colon cancer treatment. Our findings suggest that Fe3O4@ZIF-8 holds promise as a carrier for small-molecule drug adsorption and its ferromagnetic properties provide drug targeting capabilities, thereby enhancing therapeutic effects on tumors at the same drug dosage. With excellent biocompatibility, Fe3O4@ZIF-8 demonstrates potential as a drug carrier in targeted cancer chemotherapy. Our work suggests that a combination of magnetic targeting and acid-responsiveness holds great promise for advancing targeted cancer therapy in precision nanomedicine.

The ASAS Public Policy Committee (PPC) provides updates of Grand Challenges (GCs; www.asas.org/about/public-policy/asas-grand-challenges) to clearly articulate research priorities, to provide science-based information for shaping public policy, and to enhance future funding for research and education programs in animal sciences (AS). In this nexus symposium for 2021, PPC examines previous stated priorities and provides a progress report card and offers additional perspectives and recommendations for research needed to address some of the GCs continuing to face animal agriculture. Among the GC topics is growth and development, a rather broad field of inquiry focused on improving the overall growth efficiency of meat producing animals. The genesis of this discipline and its popularity grew mainly in response to heightened efforts by pharmaceutical companies to identify, develop and adopt novel new growth promotants. This included a myriad of work on the highly heralded technologies involving estrogenic and androgenic implants, somatotropin, and beta-adrenergic agonists. Because the potential application of these technologies was so broad, many disciplines within the animal sciences became involved in the process of creating knowledge around these drivers of productivity. In the process, our understanding of how tissues grow in response to these compounds, under a myriad of other conditions, and our fundamental understanding of the molecular and cellular mechanisms regulating growth and development, expanded significantly. Areas of significant expansion included but were not restricted to: satellite cell biology and myogenesis, whole body and tissue-specific protein synthesis and degradation, growth factor biology, adipogenesis, and repartitioning of nutrients throughout the body. In our quest to increase productivity and product quality, coupled with advances in scientific techniques, long-existing and emerging genetic mutations with desirable traits were studied and mechanisms undergirding their biology began to develop. Applying the most innovative tools for the detailed manipulation of cellular processes, great strides were made during this time. However, this eclectic area of investigation is perhaps more important than ever given the inevitable replacement of growth promotant technologies with new and emerging genomic technologies. Many biological research challenges lie ahead such as applications of gene editing, RNA control, and epigenetic regulation through fetal programming. This presentation will review some of the significant advances made in the growth and development area and explore where significant gains may be possible in the future.

KRAS mutations drive a quarter of cancer mortality, and most are undruggable. Several inhibitors of the MAPK pathway are FDA approved but poorly tolerated at the doses needed to adequately extinguish RAS/RAF/MAPK signaling in the tumor cell. We found that oncogenic KRAS signaling induced ferrous iron (Fe2+) accumulation early in and throughout mutant KRAS-mediated transformation. We converted an FDA-approved MEK inhibitor into a ferrous iron–activatable drug conjugate (FeADC) and achieved potent MAPK blockade in tumor cells while sparing normal tissues. This innovation allowed sustainable, effective treatment of tumor-bearing animals, with tumor-selective drug activation, producing superior systemic tolerability. Ferrous iron accumulation is an exploitable feature of KRAS transformation, and FeADCs hold promise for improving the treatment of KRAS-driven solid tumors.

The growth hormone receptor (GHR) gene in cattle is expressed as multiple GHR mRNA variants that differ in the 5′-untranslated region (5′-UTR). Three GHR mRNA 5′-UTR variants (named 1A, 1B, and 1C) were isolated in previous studies. Six additional GHR mRNA 5′-UTR variants (named 1D, 1E, 1F, 1G, 1H, and 1I) were discovered in the present study by using rapid amplification of cDNA ends (RACE). Ribonuclease protection assays (RPA) indicated that variant 1A was exclusively expressed in liver, variant 1F was expressed in liver and muscle, and most of the remaining GHR 5′-UTR variants were expressed in a variety of tissues including liver and muscle. The RPA also indicated that in liver and skeletal muscle, two important GH target tissues, the most abundant GHR mRNA 5′-UTR variants were 1A and 1B (liver) and 1B and 1C (muscle), respectively. In vitro translation assays indicated that the GHR 5′-UTR variants also differed in their effects on the translation of a luciferase reporter mRNA. Notably, the GHR 5′-UTR 1B, which is a highly expressed GHR 5′-UTR variant in vivo, strongly inhibited translation of the luciferase reporter mRNA. The observations in the present study suggest that GHR expression in the bovine may be controlled by translational mechanisms in addition to complex transcriptional mechanisms.

Transcription factors (TFs) are key regulators of gene expression during cell differentiation. Four TFs including Myf5, MyoD, MyoG and Myf6 have been identified as key myogenic regulatory factors (MYFs) that regulate gene transcription during myogenesis. Satellite cells (SCs) are the myogenic precursor cells in adult skeletal muscle. The objective of this study was to identify additional TFs that control the differentiation of bovine satellite cells. Bovine satellite cells (bSCs) were isolated from 4 crossbred steers and were initially cultured in growth medium for 12 days to expand and then in differentiation medium for 48 hours to differentiate. Chromatin immunoprecipitation followed by sequencing (ChIP-seq) was performed to identify chromatin regions marked with acetylation of histone H3 on lysine 27 (H3K27ac). This ChIP-seq analysis revealed 3,348 and 38,800 H3K27ac-associated chromatin regions in bSCs before and after differentiation, respectively. A motif enrichment analysis of the H3K27ac-marked chromatin regions from the differentiated bSCs indicated the enrichment of binding sites for the 4 MYFs and many other TFs including Fos and FosB. RNA-sequencing revealed the upregulation of Fos and FosB mRNAs in bSCs from growth to differentiation. To verify the roles of Fos and FosB in bSC differentiation, their expressions in bSCs were reduced by siRNA-induced knockdown. Based on qRT-PCR analyses, expressions of MYH2, MYH3, MYOG, and CKM mRNAs, which were selected as markers of muscle cell differentiation, were increased (P < 0.05) in bSCs from growth to differentiation, but the increases in at least three of them were reversed (P < 0.05) by Fos or FosB knockdown. Taken together, these results establish Fos and FosB as transcriptional regulators of bovine satellite cell differentiation.

Rumen epithelium plays an essential role in absorption, transport, and metabolism of short-chain fatty acids, the main products of rumen fermentation, and in preventing microbes and other potentially harmful rumen contents from entering the systemic circulation. The objective of this study was to generate an immortal rumen epithelial cell line that can be used as a convenient model of rumen epithelial cells in vitro. We isolated primary rumen epithelial cells from a steer through trypsin digestion and transduced them with lentiviruses expressing the Simian Virus (SV) 40 T antigen. We cloned the transduced cells by limiting dilution. Western blotting analysis confirmed the expression of the SV40 T antigen in two single-cell clones. Cells from one clone, named bovine rumen epithelial clone 1 (BREC1), displayed a flat and squamous morphology in culture. RNA sequencing revealed that BREC1 cells expressed many markers of epithelial cells, including keratins, the epidermal growth factor receptor, and the short-chain fatty acid transporters monocarboxylic acid transporter (MCT)1 (MCT-1) and MCT-4. RNA sequencing revealed that BREC1 cells expressed key enzymes such as 3-hydroxymethyl-3-methylglutaryl-CoA lyase and 3-hydroxy-3-methylglutaryl-CoA synthase 1 involved in ketogenesis, a unique function of rumen epithelial cells. RNA sequencing also revealed the expression of genes encoding tight junctions, desmosomes, anchoring junctions, and polarized plasma membranes, structures typical of epithelial cells, in BREC1 cells. Cell proliferation assays indicated that BREC1 cells were similar to primary rumen epithelial cells in response to insulin-like growth factor 1, insulin, and butyrate. In conclusion, BREC1 is not only a convenient but an appropriate model for studying the factors and mechanisms that control proliferation, apoptosis, differentiation, nutrient transport, metabolism, and barrier function in rumen epithelium.

Smooth muscle cells (SMCs) play an important role in physiology and production in farm animals such as pigs. Here, we report the generation of a pig SMC line. Our original objective was to establish an enteroendocrine cell line from the pig ileum epithelium through lentiviral transduction of the Simian Virus (SV) 40 large T antigen. However, an initial expression analysis of marker genes in nine cell clones revealed that none of them were enteroendocrine cells or absorptive enterocytes, goblet cells, or Paneth cells, some of the major cell types existing in the ileum epithelium. A more detailed characterization of one clone named PIC7 by RNA-seq showed that these cells expressed many of the known smooth muscle-specific or -enriched genes, including smooth muscle actin alpha 2, calponin 1, calponin 3, myosin heavy chain 11, myosin light chain kinase, smoothelin, tenascin C, transgelin, tropomyosin 1, and tropomyosin 2. Both quantitative PCR and RNA-seq analyses showed that the PIC7 cells had a high expression of mRNA for smooth muscle actin gamma 2, also known as enteric smooth muscle actin. A Western blot analysis confirmed the expression of SV40 T antigen in the PIC7 cells. An immunohistochemical analysis demonstrated the expression of smooth muscle actin alpha 2 filaments in the PIC7 cells. A collagen gel contraction assay showed that the PIC7 cells were capable of both spontaneous contraction and contraction in response to serotonin stimulation. We conclude that the PIC7 cells are derived from an enteric SMC from the pig ileum. These cells may be a useful model for studying the cellular and molecular physiology of pig enteric SMCs. Because pigs are similar to humans in anatomy and physiology, the PIC7 cells may be also used as a model for human intestinal SMCs.

Although the nature of ruminant evolution is still disputed, current theory based on physiology and genetic analysis suggests that the abomasum is the evolutionarily oldest stomach compartment, the rumen evolved some time after the abomasum, and the omasum is the evolutionarily youngest stomach compartment. In addition, there is some evidence of relaxed selective constraint in the stomach-like organ and the foregut shortly after the foregut formation event. Along with the assumption of a mean, stochastic rate of evolution, analysis of differences in genetic profiles among digestive body organs can give clues to the relationships among these organs. The presence of large numbers of uniquely expressed entries in the abomasum and rumen indicates either a period of relaxed selective constraint or greater evolutionary age. Additionally, differences in expression profiles indicate that the abomasum, rumen, and intestine are more closely related to each other, while the reticulum and omasum are more closely related to the rumen. Functional analysis using Gene Ontology (GO) categories also supports the proposed evolutionary relationships by identifying shared functions, such as muscle activity and development, lipid transport, and urea metabolism, between all sections of the digestive tract investigated.