An immunohistochemical study of lymphatic elements in the human brain.

Almost 150 papers about brain lymphatics have been published in the last 150 years. Recently, the information in these papers has been synthesized into a picture of central nervous system (CNS) "glymphatics," but the fine structure of lymphatic elements in the human brain based on imaging specific markers of lymphatic endothelium has not been described. We used LYVE1 and PDPN antibodies to visualize lymphatic marker-positive cells (LMPCs) in postmortem human brain samples, meninges, cavernous sinus (cavum trigeminale), and cranial nerves and bolstered our findings with a VEGFR3 antibody. LMPCs were present in the perivascular space, the walls of small and large arteries and veins, the media of large vessels along smooth muscle cell membranes, and the vascular adventitia. Lymphatic marker staining was detected in the pia mater, in the arachnoid, in venous sinuses, and among the layers of the dura mater. There were many LMPCs in the perineurium and endoneurium of cranial nerves. Soluble waste may move from the brain parenchyma via perivascular and paravascular routes to the closest subarachnoid space and then travel along the dura mater and/or cranial nerves. Particulate waste products travel along the laminae of the dura mater toward the jugular fossa, lamina cribrosa, and perineurium of the cranial nerves to enter the cervical lymphatics. CD3-positive T cells appear to be in close proximity to LMPCs in perivascular/perineural spaces throughout the brain. Both immunostaining and qPCR confirmed the presence of adhesion molecules in the CNS known to be involved in T cell migration.

Survey investigation of articaine use in the United States.

BACKGROUND: Local anesthesia is an essential component of dentistry, but there is limited quantifiable understanding of what techniques and local anesthetic solutions are used by practicing dentists. Use of the local anesthetic articaine has been highly debated in dentistry regarding its efficacy and risks for paresthesia. The aims of this study were to expand the knowledge of local anesthesia practices of dentists in the United States through a large-scale survey and associate potential influencing factors regarding articaine use specifically. METHODS: The 23-item survey was sent to 10,340 practicing dentists in the United States, gathering demographic data and local anesthesia approaches and concerns. Statistical analysis consisted of descriptive, bivariate, and multivariate logistic regression analyses. RESULTS: A total of 1,128 dentists completed the survey. Previous experience with articaine was reported by 97.6% of respondents, with 3.3% no longer using articaine. Sixty percent of respondents indicated using articaine for most local anesthetic injections administered. Multivariable regression analysis found those reporting to use articaine for all local anesthetic injections involving vasoconstrictors were more likely to be male (odds ratio, 1.59; P = .002) or general dentists (odds ratio, 1.63; P < .001). CONCLUSIONS: Articaine has a perceived benefit to practitioners as most respondents reported using articaine as their primary local anesthetic. A practitioner's sex and type were found to affect the profile of use of articaine. PRACTICAL IMPLICATIONS: Assembling evidenced-based local anesthesia practices would be beneficial to ensure US practitioners are more standardized in administering local anesthetics, particularly articaine, in the safest and most efficacious way.

Atto 465 Derivative Is a Nuclear Stain with Unique Excitation and Emission Spectra Useful for Multiplex Immunofluorescence Histochemistry.

Multiplex immunofluorescence (mIF) is an effective technique for the maximal visualization of multiple target proteins in situ. This powerful tool is mainly limited by the spectral overlap of the currently available synthetic fluorescent dyes. The fluorescence excitation wavelengths ranging between 405 and 488 nm are rarely used in mIF imaging and serve as a logical additional slot for a fluorescent probe. In the present study, we demonstrate that the addition of 2,3,4,5,6-pentafluoroaniline to Atto 465 NHS ester, creating Atto 465-pentafluoroaniline (Atto 465-p), generates a bright nuclear stain in the violet-blue region of the visible spectrum. This allows the 405 nm excitation and emission, classically used for nuclear counterstains, to be used for the detection of another target protein. This increases the flexibility of the mIF panel and, with appropriate staining and microscopy, enables the quantitative analysis of at least six targets in one tissue section. (J Histochem Cytochem XX: XXX-XXX, XXXX).