Mechanical stress is pervasive in egress routes of malignancy, yet the intrinsic effects of force on tumour cells remain poorly understood. Here, we demonstrate that frictional force characteristic of flow in the lymphatics stimulates YAP1 to drive cancer cell migration; whereas intensities of fluid wall shear stress (WSS) typical of venous or arterial flow inhibit taxis. YAP1, but not TAZ, is strictly required for WSS-enhanced cell movement, as blockade of YAP1, TEAD1-4 or the YAP1–TEAD interaction reduces cellular velocity to levels observed without flow. Silencing of TEAD phenocopies loss of YAP1, implicating transcriptional transactivation function in mediating force-enhanced cell migration. WSS dictates expression of a network of YAP1 effectors with executive roles in invasion, chemotaxis and adhesion downstream of the ROCK–LIMK–cofilin signalling axis. Altogether, these data implicate YAP1 as a fluid mechanosensor that functions to regulate genes that promote metastasis.

Summary Immunofluorescence (IF) is an important immunochemical technique that allows detection and localization of a wide variety of antigens in different types of tissues of various cell preparations. IF allows for excellent sensitivity and amplification of signal in comparison to immunohistochemistry, employing various microscopy techniques. There are two methods available, depending on the scope of the experiment or the specific antibodies in use: Direct (Primary) or Indirect (Secondary). Here, we describe preparation of specimens preserved in different types of media and step-by-step methods for both direct and indirect immunofluorescence staining.

Angiotensin II type 1 receptor (AT1R) antibody has been linked to poor allograft outcomes in adult kidney transplantation. However, its clinical consequences in children are unknown. To study this, we examined the relationship of AT1R antibody with clinical outcomes, biopsy findings, inflammatory cytokines, and HLA donor-specific antibodies (DSA) in a cohort of pediatric renal transplant recipients. Sixty-five patients were longitudinally monitored for AT1R antibody, HLA DSA, IL-8, TNF-α, IL-1β, IFN-γ, IL-17, and IL-6, renal dysfunction, hypertension, rejection, and allograft loss during the first two years post transplantation. AT1R antibody was positive in 38 of the 65 of children but was not associated with HLA DSA. AT1R antibody was associated with renal allograft loss (odds ratio of 13.1 [95% confidence interval 1.48-1728]), the presence of glomerulitis or arteritis, and significantly higher TNF-α, IL-1β, and IL-8 levels, but not rejection or hypertension. AT1R antibody was associated with significantly greater declines in eGFR in patients both with and without rejection. Furthermore, in patients without rejection, AT1R antibody was a significant risk factor for worsening eGFR over the two-year follow-up period. Thus, AT1R antibody is associated with vascular inflammation in the allograft, progressive decline in eGFR, and allograft loss. AT1R antibody and inflammatory cytokines may identify those at risk for renal vascular inflammation and lead to early biopsy and intervention in pediatric kidney transplantation.

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Hypercholesterolemia, the driving force of atherosclerosis, accelerates the expansion and mobilization of hematopoietic stem and progenitor cells (HSPCs). The molecular determinants connecting hypercholesterolemia with hematopoiesis are unclear. Here we report that a somite-derived pro-hematopoietic cue, AIBP, orchestrates HSPC emergence from the hemogenic endothelium, a type of specialized endothelium manifesting hematopoietic potential. Mechanistically, AIBP-mediated cholesterol efflux activates endothelial Srebp2, the master transcription factor for cholesterol biosynthesis, which in turn transactivates Notch and promotes HSPC emergence. Srebp2 inhibition impairs hypercholesterolemia-induced HSPC expansion. Srebp2 activation and Notch upregulation are associated with HSPC expansion in hypercholesterolemic human subjects. Genome-wide ChIP-seq, RNA-seq, and ATAC-seq indicate that Srebp2 trans-regulates Notch pathway genes required for hematopoiesis. Our studies outline an AIBP-regulated Srebp2-dependent paradigm for HSPC emergence in development and HPSC expansion in atherosclerotic cardiovascular disease.

Pathology is practiced by visual inspection of histochemically stained tissue slides. While the hematoxylin and eosin (H&E) stain is most commonly used, special stains can provide additional contrast to different tissue components. Here, we demonstrate the utility of supervised learning-based computational stain transformation from H&E to special stains (Masson’s Trichrome, periodic acid-Schiff and Jones silver stain) using kidney needle core biopsy tissue sections. Based on the evaluation by three renal pathologists, followed by adjudication by a fourth pathologist, we show that the generation of virtual special stains from existing H&E images improves the diagnosis of several non-neoplastic kidney diseases, sampled from 58 unique subjects (P = 0.0095). A second study found that the quality of the computationally generated special stains was statistically equivalent to those which were histochemically stained. This stain-to-stain transformation framework can improve preliminary diagnoses when additional special stains are needed, also providing significant savings in time and cost.

Mesenchymal stromal cells (MSCs) are believed to mobilize from the bone marrow in response to inflammation and injury, yet the effects of egress into the vasculature on MSC function are largely unknown. Here we show that wall shear stress (WSS) typical of fluid frictional forces present on the vascular lumen stimulates antioxidant and anti-inflammatory mediators, as well as chemokines capable of immune cell recruitment. WSS specifically promotes signaling through NFκB-COX2-prostaglandin E2 (PGE2) to suppress tumor necrosis factor-α (TNF-α) production by activated immune cells. Ex vivo conditioning of MSCs by WSS improved therapeutic efficacy in a rat model of traumatic brain injury, as evidenced by decreased apoptotic and M1-type activated microglia in the hippocampus. These results demonstrate that force provides critical cues to MSCs residing at the vascular interface which influence immunomodulatory and paracrine activity, and suggest the potential therapeutic use of force for MSC functional enhancement.

Diaz et al. show that biochemical forces induced by blood flow promote the development of hematopoietic cells at early embryonic stages via induction of prostaglandin E2 and signaling pathways involved in hematopoiesis.