Establishing thresholds for cytokine storm and defining their relationship to disease severity in respiratory viral infections.

Previous studies have identified cytokines associated with respiratory virus infection illness outcome. However, few studies have included comprehensive cytokine panels, longitudinal analyses, and/or simultaneous assessment across the severity spectrum. This, coupled with subjective definitions of cytokine storm syndrome (CSS), have contributed to inconsistent findings of cytokine signatures, particularly with COVID severity. Here, we measured 38 plasma cytokines and compared profiles in healthy, SARS-CoV-2 infected, and multisystem inflammatory syndrome in children (MIS-C) patients (n = 169). Infected patients spanned the severity spectrum and were classified as Asymptomatic, Mild, Moderate or Severe. Our results showed acute cytokine profiles and longitudinal dynamics of IL1Ra, IL10, MIP1b, and IP10 can differentiate COVID severity groups. Only 4% of acutely infected patients exhibited hypercytokinemia. Of these subjects, 3 were Mild, 3 Moderate, and 1 Severe, highlighting the lack of association between CSS and COVID severity. Additionally, we identified IL1Ra and TNFa as potential biomarkers for patients at high risk for long COVID. Lastly, we compare hypercytokinemia profiles across COVID and influenza patients and show distinct elevated cytokine signatures, wherein influenza induces the most elevated cytokine profile. Together, these results identify key analytes that, if obtained at early time points, can predict COVID illness outcome and/or risk of complications, and provide novel insight for improving the conceptual framework of hypercytokinemia, wherein CSS is a subgroup that requires concomitant severe clinical manifestations, and including a list of cytokines that can distinguish between subtypes of hypercytokinemia.

The Use of Left Ventricular Assist Devices in End-Stage Heart Failure.

Heart failure is one of the most common diagnoses in patients requiring multiple hospitalizations. The use of mechanical circulatory support (both temporary and durable), especially left ventricular assist devices (LVADs), in this patient population has evolved over the last decade. There are thousands of durable LVADs implanted in the United States annually as bridge to transplantation, destination therapy, or bridge to recovery. LVAD therapy, just like cardiac transplantation, takes a multidisciplinary team approach to achieve success. Appropriate patient selection is the key to good clinical outcomes. As the technology of LVAD therapy continues to evolve, it will be made available to an increasing number of patients in an attempt to improve their morbidity and mortality.

Deregulation of cofactor of BRCA1 expression in breast cancer cells.

Cofactor of BRCA1 (COBRA1) is an integral component of the human negative elongation factor (NELF), a four-subunit protein complex that inhibits transcription elongation. Previous in vivo work indicates that COBRA1 and the rest of the NELF complex repress estrogen-dependent transcription and the growth of breast cancer cells. In light of the COBRA1 function in breast cancer-related gene expression, we sought to examine regulation of COBRA1 expression in both established breast cancer cell lines and breast carcinoma tissues. We found that COBRA1 expression was inversely correlated with breast cancer progression, as tumor samples of patients who had distant metastasis and local recurrence expressed very low levels of COBRA1 mRNA when compared to those who were disease free for over 10 years (P = 0.0065 and 0.0081, respectively). Using both breast and prostate cancer cell lines, we also explored the possible mechanisms by which COBRA1 expression is regulated. Our results indicate that the protein abundance of COBRA1 and the other NELF subunits are mutually influenced in a tightly coordinated fashion. Small interfering RNA (siRNA) that targeted at one NELF subunit dampened the protein levels of all four subunits. Conversely, ectopic expression of COBRA1 in the knockdown cells partially rescues the co-depletion of the NELF subunits. In addition, our study suggests that a post-transcriptional, proteasome-independent mechanism is involved in the interdependent regulation of the NELF abundance. Furthermore, a lack of COBRA1 expression in breast carcinoma may serve as a useful indicator for poor prognosis.

Cofactor of BRCA1 modulates androgen-dependent transcription and alternative splicing.

Transcriptional activity of nuclear receptors (NRs) is influenced by a large number of coregulators that exert their actions predominantly at the transcription initiation step. Unlike most well-characterized NR coregulators, cofactor of BRCA1 (COBRA1), a subunit of the negative elongation factor (NELF), binds to estrogen receptor alpha (ERalpha) and modulates estrogen-dependent transcription by impeding the movement of RNA polymerase II (RNAPII) during the transcription elongation stage. Here we show that, in addition to ERalpha, COBRA1 also displays various degrees of affinity for several other NRs. In particular, COBRA1 binds strongly to androgen receptor (AR) via its ligand-binding domain (LBD). Small hairpin RNA (shRNA)-mediated reduction of endogenous COBRA1 enhances androgen-mediated transcription. The effect of COBRA1 knockdown can be rescued by a silent mutant COBRA1 that is refractory to the shRNA action. Using a reporter assay for alternative splicing, we also provide evidence for a role of COBRA1 in influencing the exon skipping/inclusion of nascent transcripts produced from an androgen-dependent promoter. These findings suggest that COBRA1 may coordinate multiple steps in ligand-dependent gene expression, which in turn ensures both the quantity and quality of hormone-stimulated gene products.

Attenuation of estrogen receptor alpha-mediated transcription through estrogen-stimulated recruitment of a negative elongation factor.

Estrogen receptor alpha (ERalpha) signaling is paramount for normal mammary gland development and function and the repression of breast cancer. ERalpha function in gene regulation is mediated by a number of coactivators and corepressors, most of which are known to modify chromatin structure and/or influence the assembly of the regulatory complexes at the level of transcription initiation. Here we describe a novel mechanism of attenuating the ERalpha activity. We show that cofactor of BRCA1 (COBRA1), an integral subunit of the human negative elongation factor (NELF), directly binds to ERalpha and represses ERalpha-mediated transcription. Reduction of the endogenous NELF proteins in breast cancer cells using small interfering RNA results in elevated ERalpha-mediated transcription and enhanced cell proliferation. Chromatin immunoprecipitation reveals that recruitment of COBRA1 and the other NELF subunits to endogenous ERalpha-responsive promoters is greatly stimulated upon estrogen treatment. Interestingly, COBRA1 does not affect the estrogen-dependent assembly of transcription regulatory complexes at the ERalpha-regulated promoters. Rather, it causes RNA polymerase II (RNAPII) to pause at the promoter-proximal region, which is consistent with its in vitro biochemical activity. Therefore, our in vivo work defines the first corepressor of nuclear receptors that modulates ERalpha-dependent gene expression by stalling RNAPII. We suggest that this new level of regulation may be important to control the duration and magnitude of a rapid and reversible hormonal response.