Thrombospondin-1 (TSP1) is a matricellular protein with many important roles in mediating carcinogenesis, fibrosis, leukocyte recruitment, and metabolism. We have previously shown a role of diet in the absence of TSP1 in liver metabolism in the context of a colorectal cancer model. However, the metabolic implications of TSP1 regulation by diet in the liver metabolism are currently understudied. Therefore Discrete correlation summation (DCS) was used to re-interrogate data and determine the metabolic alterations of TSP1 deficiency in the liver, providing new insights into the role of TSP1 in liver injury and the progression of liver pathologies such as nonalcoholic fatty liver disease (NAFLD). DCS analysis provides a straightforward approach to rank covariance and data clustering when analyzing complex data sets. Using this approach, our previous liver metabolite data was re-analyzed by comparing wild-type (WT) and Thrombospondin-1 null (Thbs1^−/−) mice, identifying changes driven by genotype and diet. Principal component analysis showed clustering of animals by genotype regardless of diet, indicating that TSP1 deficiency alters metabolite handling in the liver. High-fat diet consumption significantly altered over 150 metabolites in the Thbs1^−/− livers versus approximately 90 in the wild-type livers, most involved in amino acid metabolism. The absence of Thbs1 differentially regulated tryptophan and tricarboxylic acid cycle metabolites implicated in the progression of NAFLD. Overall, the lack of Thbs1 caused a significant shift in liver metabolism with potential implications for liver injury and the progression of NAFLD.

Muscadine grapes are abundant in dietary polyphenols, but their effect on hypertension-induced cardiac damage is limited. This study assessed whether a muscadine grape skin/seed extract supplement (MGES) prevents hypertension-induced cardiac damage and oxidative stress. Male Sprague Dawley rats were treated for four weeks with drinking water, angiotensin II (Ang II) to induce hypertension, MGES, or both Ang II and MGES. Cardiac function assessed by echocardiography showed that Ang II increased systolic blood pressure while MGES alone or in combination with Ang II had no effect. Ang II increased E/e′, an indicator of left ventricular filling pressure and diastolic dysfunction, by 41% compared to Control and co-treatment with MGES prevented the Ang II-mediated increase, suggesting that the extract attenuated hypertension-induced diastolic function. Ang II infusion increased urinary 8-hydroxy-2′-deoxyguanosine and cardiac 4-hydroxynonenal and malondialdehyde, which were prevented by the extract. The antioxidant enzymes catalase and superoxide dismutase 1 activity and mRNA were increased significantly in animals treated with MGES alone or in combination with Ang II, suggesting that the extract upregulates oxidative stress defense mechanisms in cardiac tissue. Thus, MGES may serve as a medical food to protect the heart from hypertension-induced diastolic dysfunction caused in part by excessive reactive oxygen species production.

Most women diagnosed with breast cancer (BC) have estrogen receptor alpha-positive (ER+) disease. The current mouse models of ER+ BC often rely on exogenous estrogen to encourage metastasis, which modifies the immune system and the function of some tissues like bone. Other studies use genetically modified or immunocompromised mouse strains, which do not accurately replicate the clinical disease. To create a model of antiestrogen responsive BC with spontaneous metastasis, we developed a mouse model of 4T1.2 triple-negative (TN) breast cancer with virally transduced ER expression that metastasizes spontaneously without exogenous estrogen stimulation and is responsive to antiestrogen drugs. Our mouse model exhibited upregulated ER-responsive genes and multi-organ metastasis without exogenous estrogen administration. Additionally, we developed a second TN BC cell line, E0771/bone, to express ER, and while it expressed ER-responsive genes, it lacked spontaneous metastasis to clinically important tissues. Following antiestrogen treatment (tamoxifen, ICI 182,780, or vehicle control), 4T1.2- and E0771/bone-derived tumor volumes and weights were significantly decreased, exemplifying antiestrogen responsivity in both cell lines. This 4T1.2 tumor model, which expresses the estrogen receptor, metastasizes spontaneously, and responds to antiestrogen treatment, will allow for further investigation into the biology and potential treatment of metastasis.

Background: Triple-negative breast cancer requires new treatment strategies due to poor responses to current therapies. While myeloid SIRPα mediates immunosuppression, its cancer intrinsic role remains poorly understood. Methods: Human breast cancer scRNAseq profiles were used to examine SIRPα expression across different cell populations and subtypes. TNBC brain-tropic cells were injected into the mouse mammary fat pad for the orthotopic tumor model and intracardiac-injected for brain metastasis models. Bulk RNA sequencing was used to determine SIRPα-regulated pathway. Stably SIRPα overexpressed and knockout TNBC cell lines were established to determine SIRPα intracellular regulation. Digital spatial profiling was utilized to investigate the orthotopic and brain metastasis tumor immune microenvironment. Results: Human single-cell data showed that SIRPα levels increased in malignant TNBC epithelial cells. We observed that SIRPα is upregulated in patient breast-to-brain metastatic lesions. SIRPα is overexpressed in TNBC brain-tropic cells compared to parental cells. Bulk RNA-Seq showed that targeting SIRPα affects genes involved in mitochondrial dynamics, and that SIRPα upregulates mitochondrial fission and induces metastasis through the SHP2/Erk/Drp1 signaling pathway. In vivo, overexpression of SIRPα in cancer cells significantly increases TNBC systemic metastasis. Next, spatial proteomics revealed changes in the immune microenvironment associated with the SIRPα-regulated ECM protein fibronectin. Fibronectin induces microglial tolerance by impairing inflammatory signaling and metabolic reprogramming, allowing cancer to escape microglial immunosurveillance. Most importantly, SIRPα inhibition reduced TNBC brain metastatic lesions in mouse metastasis models. Conclusion: Cancer-intrinsic SIRPα promotes TNBC brain metastasis through increased mitochondria fission and triggering microglia tolerance, and targeting SIRPα reduces brain metastasis.

We present a model of directed investigation of KEGG pathway analysis, beginning with discrete correlate summation (DCS) clustering of Swiss-Prot sets. Dopamine and angiotensin related proteins define the boundary for pathway analyses upon a network of proteins also associated with a compositely defined matrix of Swiss-Prot keywords to provide a framework for their biological indices (Figure 1).

Objective: Preeclampsia increases long-term cardiovascular, cerebrovascular, and cognitive risk but the underlying mechanisms remain unclear. We examined cerebrovascular function, cognitive performance, and placenta-derived extracellular-vesicle (PD-EV) microRNA profiles in a transgenic rat model with preeclamptic features (TGA-PE) at 2 months after delivery (2M). Methods: Middle cerebral artery (MCA) hemodynamics were assessed by transcranial Doppler (TCD), and cerebral oxygenation was measured by photoacoustic imaging (PAI) in TGA-PE and Sprague-Dawley control rats. Blood pressure was measured using a tail-cuff method. Touchscreen-based cognitive tasks evaluated learning and reward-directed responding, while histological assessment evaluated MCA structure. MicroRNA sequencing was performed on circulating PD-EV, and miRNA target networks were analyzed using KEGG-based NEST-Scoring (Node, Edge, System, Topology). Validation of miR-363-3p was performed in PD-EV, plasma, non-PD-EV, and human brain endothelial cells (HBEC-5i). Results: TGA-PE rats were hypertensive and exhibited lower MCA resistance and pulsatility indexes with unchanged blood flow velocities. Cognitive testing revealed impaired associative learning and reduced motivation to respond on a progressive-ratio schedule. Temporal brain oxygen saturation was higher in TGA-PE rats with unchanged total hemoglobin, lower brain HIF-2α, and unchanged VEGF levels. MCA elastin content was reduced in TGA-PE rats without changes in total collagen. PD-EV sequencing identified 29 differentially expressed miRNAs, including reduced miR-363-3p in non-placental EVs and plasma at 2M. Inhibition of miR-363-3p in human cerebral endothelial cell line HBEC-5i upregulated NOX4, p21, and β-galactosidase levels. NEST-Scoring highlighted pathways involving MAPK, Ras, TNF, focal adhesion, cytoskeletal regulation, and cellular senescence. Conclusions: The exposure to experimental preeclampsia resulted in altered MCA flow resistance, cerebral oxygenation, vascular structure and behavioral performance. Although direct mechanistic studies are needed to establish the role of miRNA363-3p in neurovascular health, the increased endothelial oxidative stress and senescence-associated markers in HBEC-5i cells in response to miRNA363-3p inhibition suggest a potential protective role for miRNA363-3p. A novel NEST-Scoring analysis also identified potential pathways for future mechanistic investigations. Overall, the TGA-PE model provides a useful experimental system for investigating candidate mechanisms that may contribute to long-term maternal neurovascular changes following hypertensive pregnancy and sequelae of preeclamptic pathophysiology.