Semi-automated protocol to quantify and characterize fluorescent three-dimensional vascular images.

The microvasculature facilitates gas exchange, provides nutrients to cells, and regulates blood flow in response to stimuli. Vascular abnormalities are an indicator of pathology for various conditions, such as compromised vessel integrity in small vessel disease and angiogenesis in tumors. Traditional immunohistochemistry enables the visualization of tissue cross-sections containing exogenously labeled vasculature. Although this approach can be utilized to quantify vascular changes within small fields of view, it is not a practical way to study the vasculature on the scale of whole organs. Three-dimensional (3D) imaging presents a more appropriate method to visualize the vascular architecture in tissue. Here we describe the complete protocol that we use to characterize the vasculature of different organs in mice encompassing the methods to fluorescently label vessels, optically clear tissue, collect 3D vascular images, and quantify these vascular images with a semi-automated approach. To validate the automated segmentation of vascular images, one user manually segmented one hundred random regions of interest across different vascular images. The automated segmentation results had an average sensitivity of 83±11% and an average specificity of 91±6% when compared to manual segmentation. Applying this procedure of image analysis presents a method to reliably quantify and characterize vascular networks in a timely fashion. This procedure is also applicable to other methods of tissue clearing and vascular labels that generate 3D images of microvasculature.

Pattern Completion and Rate Remapping in Retrosplenial Cortex.

Principles governing the encoding, storage, and updating of memories in cortical networks are poorly understood. In retrosplenial cortex (RSC), cells respond to the animal's position as it navigates a real or virtual (VR) linear track. Position correlated cells (PCCs) in RSC require an intact hippocampus to form. To examine whether PCCs undergo pattern completion and remapping like hippocampal cells, neuronal activity in RSC or CA1 was recorded using two-photon calcium imaging in mice running on VR tracks. RSC and CA1 PCC activity underwent global and rate remapping depending on the degree of change to familiar environments. The formation of position correlated fields in both regions required stability across laps; however, once formed, PCCs became robust to object destabilization, indicating pattern completion of the previously formed memory. Thus, memory and remapping properties were conserved between RSC and CA1, suggesting that these functional properties are transmitted to cortex to support memory functions.

Distinct SARS-CoV-2 antibody reactivity patterns elicited by natural infection and mRNA vaccination.

We analyzed data from two ongoing COVID-19 longitudinal serological surveys in Orange County, CA., between April 2020 and March 2021. A total of 8476 finger stick blood specimens were collected before and after a vaccination campaign. IgG levels were determined using a multiplex antigen microarray containing antigens from SARS-CoV-2, SARS, MERS, Common CoV, and Influenza. Twenty-six percent of specimens from unvaccinated Orange County residents in December 2020 were SARS-CoV-2 seropositive; out of 852 seropositive individuals 77 had symptoms and 9 sought medical care. The antibody response was predominantly against nucleocapsid (NP), full length, and S2 domain of spike. Anti-receptor binding domain (RBD) reactivity was low and not cross-reactive against SARS S1 or SARS RBD. A vaccination campaign at the University of California Irvine Medical Center (UCIMC) started on December, 2020 and 6724 healthcare workers were vaccinated within 3 weeks. Seroprevalence increased from 13% pre-vaccination to 79% post-vaccination in January, 93% in February, and 99% in March. mRNA vaccination induced higher antibody levels than natural exposure, especially against the RBD domain and cross-reactivity against SARS RBD and S1 was observed. Nucleocapsid protein antibodies can be used to distinguish vaccinees to classify pre-exposure to SARS-CoV-2 Previously infected individuals developed higher antibody titers to the vaccine than non pre-exposed individuals. Hospitalized patients in intensive care with severe disease reach significantly higher antibody levels than mild cases, but lower antibody levels compared to the vaccine. These results indicate that mRNA vaccination rapidly induces a much stronger and broader antibody response than SARS-CoV-2 infection.

Considerations for Cannabis Use to Treat Pain in Sickle Cell Disease.

Pain in Sickle Cell Disease (SCD) is a major comorbidity and unique with acute pain due to recurrent and episodic vaso-occlusive crises as well as chronic pain, which can span an individual's entire life. Opioids are the mainstay treatment for pain in SCD. Due to recent health crises raised by adverse effects including deaths from opioid use, pain management in SCD is adversely affected. Cannabis and its products are most widely used for pain in multiple conditions and also by patients with SCD on their own. With the availability of "Medical Cannabis" and approval to use cannabis as medicine across majority of States in the United States as well as over-the-counter preparations, cannabis products are being used increasingly for SCD. The reliability of many of these products remains questionable, which poses a major health risk to the vulnerable individuals seeking pain relief. Therefore, this review provides up to date insights into available categories of cannabis-based treatment strategies, their mechanism of action and pre-clinical and clinical outcomes in SCD. It provides evidence for the benefits and risks of cannabis use in SCD and cautions about the unreliable and unvalidated products that may be adulterated with life-threatening non-cannabis compounds.