Objective To characterize the functional impairments of a cohort of patients undergoing inpatient rehabilitation after surviving severe COVID-19 illness, in order to better understand the ongoing needs of this patient population. Methods This study consisted of a retrospective chart review of consecutive patients hospitalized for COVID-19 and admitted to a regional inpatient rehabilitation hospital from April 29th to May 22nd, 2020. Patient demographics, clinical characteristics and complications from acute hospitalization were examined. Measures of fall risk (Berg Balance Scale), endurance (6 Minute Walk Test), gait speed (10 Meter Walk Test), mobility (transfer and ambulation independence), cognition, speech and swallowing (American Speech and Hearing Association National Outcomes Measurement System Functional Communication Measures) were assessed at rehabilitation admission and discharge. Results The study population included 29 patients and was 70% male, 58.6% white and with a mean age of 59.5. The mean length of acute hospitalization was 32.2 days with a mean of 18.7 days intubated. Patients spent a mean of 16.7 days in inpatient rehabilitation and 90% were discharged home. Patients demonstrated significant improvement from admission to discharge in measures of fall risk, endurance, gait speed, mobility, cognition, speech and swallowing, (p< 0.05). At discharge, a significant portion of the population continued to deficits in cognition (attention 37%; memory 28%; problem solving 28%), balance (55%) and gait speed (97%). Conclusion Patients admitted to inpatient rehabilitation after hospitalization with COVID-19 demonstrated deficits in mobility, cognition, speech and swallowing at admission and improved significantly in all of these domains by discharge. However, a significant number of patients exhibited residual deficits at discharge highlighting the post-acute care needs of this patient population.

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The cDNA for a novel truncated progesterone receptor (PR-M) was previously cloned from human adipose and aortic cDNA libraries. The predicted protein sequence contains 16 unique N-terminal amino acids, encoded by a sequence in the distal third intron of the progesterone receptor PR gene, followed by the same amino acid sequence encoded by exons 4 through 8 of the nuclear PR. Thus, PR-M lacks the N terminus A/B domains and the C domain for DNA binding, whereas containing the hinge and hormone-binding domains. In this report, we have localized PR-M to mitochondria using immunofluorescent localization of a PR-M-green fluorescent protein (GFP) fusion protein and in Western blot analyses of purified human heart mitochondrial protein. Removal of the putative N-terminal mitochondrial localization signal obviated association of PR-M with mitochondria, whereas addition of the mitochondrial localization signal to green fluorescent protein resulted in mitochondrial localization. Immunoelectron microscopy and Western blot analysis after mitochondrial fractionation identified PR-M in the outer mitochondrial membrane. Antibody specificity was shown by mass spectrometry identification of a PR peptide in a mitochondrial membrane protein isolation. Cell models of overexpression and gene silencing of PR-M demonstrated a progestin-induced increase in mitochondrial membrane potential and an increase in oxygen consumption consistent with an increase in cellular respiration. This is the first example of a truncated steroid receptor, lacking a DNA-binding domain that localizes to the mitochondrion and initiates direct non-nuclear progesterone action. We hypothesize that progesterone may directly affect cellular energy production to meet the increased metabolic demands of pregnancy.

Observational studies demonstrate that estradiol and progesterone affect vasoreactivity. In animal studies, progesterone treatment causes immediate relaxation of precontracted arteries with inhibition of calcium influx in vascular endothelial and smooth muscle cells, suggesting a non-genomic mechanism of action. In this study we investigated the presence of novel membrane-bound progesterone receptors in human aortic endothelial cells and correlated the expression with cell-cycle stage. Western blotting analysis with an antibody directed to the hormone-binding domain of the classic progesterone receptors shows predominant bands at 100 and 60 kD, whereas analysis with an antibody to the DNA-binding region shows only the 100-kD band. In contrast, classic nuclear progesterone receptors B and A are identified at 116 and 94 kD in similarly processed T47D cells. Both novel bands localize to the membrane fraction after differential centrifugation. Plasma membrane-bound progesterone receptor was further shown with immunofluorescent antibody and ligand-binding studies in a small percentage of human aortic endothelial cells. Fluorescent activated cell sorting demonstrated that approximately 8% of the human aortic endothelial cells expressed a plasma membrane progesterone receptor and that a greater percentage of the expressing cells were in the G2/M-phase of the cell cycle. Treatment with progesterone conjugated to BSA did not show any significant cell-cycle changes. Plasma membrane-bound progesterone receptor in vascular endothelial cells may regulate the non-genomic actions of progesterone, and expression of the receptor appears to vary with cell cycle stage.

In many chronic neurodegenerative diseases including Frontotemporal Dementia and Alzheimer's disease (AD), microglial activation is suggested to be involved in pathogenesis or disease progression. Activated microglia secrete a variety of cytokines, including interleukin-1beta, interleukin-6, and tumor necrosis factor as well as reactive oxygen and nitrogen species (ROS/RNS). ROS and RNS contribute to alterations in neuronal glucose uptake, inhibition of mitochondrial enzymes, a decrease in mitochondrial membrane potential, impaired axonal transport, and synaptic signaling. In addition, ROS act as signaling molecules in pro-inflammatory redox-active signal transduction pathways. To establish a high throughput screening system for anti-inflammatory and neuroprotective compounds, we have constructed an "Enhanced Green Fluorescent protein" (EGFP) expressing neuronal cell line and set up a murine microglia/neuron co-culture system with these EGFP expressing neuronal cells. We show that microglia activation leads to neuronal cell death, which can be conveniently measured by loss of neuronal EGFP fluorescence. Moreover, we used this system to test selected polyphenolic compounds for their ability to downregulate inflammatory markers and to protect neurons against microglial insult. We suggest that this system might allow accelerated drug discovery for the treatment of inflammation-mediated neurodegenerative diseases.

SummaryExtracellular ATP activates purinergic (P 2 ) receptors with an increase in intracellular calcium and phosphorylation of MAPK. In this study we have investigated the effect of progesterone/progestin on ATP-induced calcium mobilization and phosphorylation of the kinase ERK in the T47D-Y breast cancer cell line that exhibits no detectable nuclear progesterone receptor expression. Brief pretreatment with progesterone/progestin results in a dose dependent inhibition of ATP-induced intracellular calcium mobilization, and inhibition of ERK phosphorylation. Response to a cell impermeable ligand and inhibition of the response by an inactivating antibody suggests a mechanism of action at the plasma membrane. These results in T47D-Y cells strongly suggest that progesterone can act in a rapid non-nuclear manner to inhibit extracellular ATP effects on intracellular calcium mobilization and ERK activation. This research provides an example of progesterone action in a breast cancer cell line lacking expression of the classical nuclear progesterone receptors.

These studies support the expression of a novel, truncated PR (PR-M) in a breast cancer cell line known to lack expression of genomic PR. This observation raises the possibility of progesterone action in breast cancer cells classically considered nonresponsive due to lack of genomic PR expression.

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