Multiplex Gene Tagging with CRISPR-Cas9 for Live-Cell Microscopy and Application to Study the Role of SARS-CoV-2 Proteins in Autophagy, Mitochondrial Dynamics, and Cell Growth.

The lack of efficient tools to label multiple endogenous targets in cell lines without staining or fixation has limited our ability to track physiological and pathological changes in cells over time via live-cell studies. Here, we outline the FAST-HDR vector system to be used in combination with CRISPR-Cas9 to allow visual live-cell studies of up to three endogenous proteins within the same cell line. Our approach utilizes a novel set of advanced donor plasmids for homology-directed repair and a streamlined workflow optimized for microscopy-based cell screening to create genetically modified cell lines that do not require staining or fixation to accommodate microscopy-based studies.

Measuring intracellular calcium signaling in murine NK cells by flow cytometry.

This chapter describes a method by which activating receptor-mediated calcium signaling can be measured in individual murine NK cells using a flow cytometer fitted with a UV laser. One major advantage of this method is that the calcium response of the minority NK cell population and even smaller NK cell subpopulations can be measured simultaneously from a mixture of freshly prepared total splenocytes without resorting to prior cell sorting or expansion in culture. Briefly, cells are harvested and stained to mark the populations of interest, then loaded with indo-1 AM dye and analyzed on the flow cytometer.

Vibrio parahaemolyticus growth under low-iron conditions and survival under high-magnesium conditions.

Since 1996, Vibrio parahaemolyticus serotype O3:K6 and closely related strains have been associated with an increased incidence of V. parahaemolyticus gastroenteritis worldwide, suggesting the emergence of strains with enhanced abilities to cause disease. One hypothesis for the recent emergence of V.

Discrimination of men with prostate cancer from those with benign disease by measurements of human glandular kallikrein 2 (HK2) in serum.

Purpose: To investigate the clinical value of measuring human glandular kallikrein 2 (hK2) compared with free and total prostate specific antigen (PSA-F and PSA-T) in serum from patients with prostate disease.

Materials And Methods: Serum from healthy controls, from men with benign prostate hyperplasia (BPH), clinically localized prostate cancer (PCa), and advanced PCa were analyzed for hK2 (using an in-house-research assay with detection limit of 0.05 ng.

Prostate specific antigen: a decade of discovery--what we have learned and where we are going.

Purpose: Many advances have occurred during the last decade in the clinical use of prostate specific antigen (PSA) for detecting, staging and monitoring prostate cancer. We review the clinical usefulness and limitations of serum PSA as a tumor marker of prostate cancer.

Materials And Methods: The English language literature was reviewed with respect to the major contributions and limitations of PSA in present clinical practice.

Have complication rates decreased after treatment for localized prostate cancer?

Purpose: The American Urological Association Prostate Cancer Clinical Guidelines Panel reviewed 12,501 publications on prostate cancer from 1955 to 1992 to determine whether the complication rates of external beam radiation therapy, interstitial radiotherapy and radical prostatectomy have decreased.

Materials And Methods: Complications reported in at least 6 series, study duration and sample sizes were extracted. Year specific study weighted mean patient ages and complication rates were computed.

Reference ranges for serum prostate-specific antigen in black and white men without cancer.

Objectives: To determine the age- and race-specific prostate-specific antigen (PSA) distributions in healthy men in central South Carolina and to compare these to data from other studies.

Methods: Two thousand ninety-two black men aged 40 to 69 years and white men aged 50 to 69 years from the general population in 11 counties of central South Carolina participated in a prostate cancer educational program. Seventy-two percent of the participants were black-about double the proportion in the general population-and 63% of the men (1319 of 2092) subsequently obtained a PSA determination from their own physician.

Screening digital rectal examination and prostate cancer mortality: a population-based case-control study.

Objectives: Although digital rectal examination (DRE) for the detection of prostate cancer has been recommended by many professional groups and has become part of the general physical examination, no randomized clinical trial has demonstrated the efficacy or effectiveness of this practice. We conducted a population-based case-control study to evaluate the association between DRE and prostate cancer mortality.

Methods: With the resources of the Rochester Epidemiology Project, all 173 men who died of prostate cancer in Olmsted County from 1976 to 1991, who were resident at the time of diagnosis, were identified.

Understanding artificial neural networks and exploring their potential applications for the practicing urologist.

Artificial neural networks (ANNs) are complex mathematical models that are distantly based on the human neuronal structure. They are capable of modeling elaborate biologic systems without making assumptions based on statistical distributions. Preliminary work has been reported on their application in urology.

Prostate cancer screening and beliefs about treatment efficacy: a national survey of primary care physicians and urologists.

Purpose: To describe practice patterns and beliefs of primary care physicians and urologists regarding early detection and treatment of prostate cancer.

Subjects And Methods: National probability samples of primary care physicians (n=444) and urologists (n=394) completed mail survey instruments in 1995. Physicians were asked about their use of prostate-specific antigen (PSA) testing for men of different ages and their beliefs about the value of radical prostatectomy, external-beam radiation therapy, and watchful waiting for men with differing life expectancies.

Transurethral needle ablation of the prostate: a minimally invasive treatment for symptomatic benign prostatic hyperplasia.

Transurethral needle ablation of the prostate, a relatively new minimally invasive treatment modality for patients with bladder outlet obstruction attributable to an enlarged prostate gland, has undergone extensive evaluation by numerous investigators worldwide. The results to date indicate that needle ablation is safe and effective for relieving symptoms in patients with benign prostatic hyperplasia, and the effect has been demonstrated to be durable for at least 2 years. Nevertheless, additional investigations with longer follow-up data are needed to address the important issues of extended durability (5 to 10 years) and biophysiologic mechanism of action.

Alpha-blockade therapy for benign prostatic hyperplasia: from a nonselective to a more selective alpha1A-adrenergic antagonist.

Benign prostatic hyperplasia (BPH) is very common in older men, causing symptoms that can markedly impair quality of life. Surgical treatment, typically transurethral resection of the prostate (TURP), is highly effective but can be costly and is associated with the risk for significant morbidity. Medical treatments for BPH are targeted toward reducing bladder outlet obstruction either by androgen blockade to reduce prostatic volume or alpha-adrenergic blockade to relax the smooth muscle tone of the prostate.

Biologic variability of prostate-specific antigen and its usefulness as a marker for prostate cancer: effects of finasteride. Finasteride PSA Study Group.

Objectives: The effects of finasteride on prostate-specific antigen (PSA) variability and usefulness in prostate cancer detection were examined.

Methods: Percent change and crossover of PSA levels between the low (1.0 to 3.

Serum percent-free PSA does not predict extraprostatic spread of prostate cancer.

Percent-free prostate-specific antigen (proportion of free prostate-specific antigen [PSA] to total PSA) has been shown recently in studies on frozen serum samples to be more useful than total PSA alone in distinguishing prostate cancer from benign conditions of the prostate gland. The primary purpose of our study was to determine whether percent-free PSA could predict extraprostatic spread of prostate cancer. We also sought to evaluate the freeze-thaw stability of free PSA.

A prospective, randomized 1-year clinical trial comparing transurethral needle ablation to transurethral resection of the prostate for the treatment of symptomatic benign prostatic hyperplasia.

Purpose: We assess the 1-year efficacy and safety of transurethral needle ablation of the prostate compared to transurethral resection of the prostate for the treatment of symptomatic benign prostatic hyperplasia (BPH).

Materials And Methods: A prospective, randomized clinical trial of 121 men 50 years old or older with symptomatic BPH was performed at 7 medical centers across the United States. Of the men 65 (54%) were treated with transurethral needle ablation of the prostate and 56 (46%) underwent transurethral resection of the prostate.

Benign prostatic hyperplasia: use of drug therapy in primary care.

Age and normal male androgenic function are the two most well-established risk factors for benign prostatic hyperplasia. Clinical manifestations of BPH may range from minimally bothersome symptoms to urinary retention and renal failure. Digital rectal examination, serum PSA, urinalysis, serum creatinine, and the AUA symptom score are recommended for the initial evaluation.

Transurethral needle ablation for benign prostatic hyperplasia: 12-month results of a prospective, multicenter U.S. study.

Objectives: To report the safety and efficacy of the transurethral needle ablation (TUNA) procedure for the treatment of clinical benign prostatic hyperplasia (BPH).

Methods: One hundred thirty patients with BPH were enrolled in two identical protocols and treated by the TUNA procedure. Entry criteria included an American Urological Association symptom index (AUA SI) of 13 points or higher and a peak flow rate of 12 mL/s or less.

A model for the number of cores per prostate biopsy based on patient age and prostate gland volume.

Purpose: The systematic sextant biopsy is currently the gold standard for the tissue diagnosis of prostate cancer. However, it is unknown whether this 6 core approach provides optimal sampling of all prostate glands in men of all ages. The goal of the current study was to determine the appropriate number of cores per prostate biopsy based on patient age and prostate gland volume.

Serial prostate-specific antigen measurements in men with clinically benign prostatic hyperplasia during a 12-month placebo-controlled study with terazosin. HYCAT Investigator Group. Hytrin Community Assessment Trial.

Objectives: To prospectively analyze whether the treatment of men with clinically benign prostatic hyperplasia (BPH) with alpha blocking agents affects the serum prostate-specific antigen (PSA) levels, and to determine the magnitude of such effect.

Methods: Serial PSA measurements were performed using the Abbott IMx assay over 1 year in 134 men over the age of 55 years participating in the Hytrin Community Assessment Trial (HYCAT). HYCAT is a 1-year, randomized, placebo-controlled, double-blinded study of the alpha1-adrenergic antagonist terazosin.

Update on the appropriate staging evaluation for newly diagnosed prostate cancer.

Purpose: Prostate cancer clinical staging methods and decision support tools were reviewed to assess their accuracy to predict pathological staging results and determine what comprises an appropriate clinical staging evaluation.

Materials And Methods: The MEDLINE data base was searched and 238 abstracts were obtained. Data were extracted from 142 articles that evaluated the preoperative accuracy of digital rectal examination, prostate specific antigen, prostatic acid phosphatase, systematic biopsy parameters (including Gleason scoring), seminal vesicle biopsy, various imaging studies and pelvic lymphadenectomy versus pathological staging results.

A nationwide survey of practicing urologists: current management of benign prostatic hyperplasia and clinically localized prostate cancer.

Purpose: Our aim was to define the spectrum of urological care for benign prostatic hyperplasia (BPH) and clinically localized prostate cancer.

Materials And Methods: In 1995 a random sample of 394 American urologists was surveyed with a response rate of 67%.

Results: Respondents reported seeing a median of 240 BPH patients during the preceding 12 months, and they had prescribed alpha-blockers for 70 and finasteride for 15.

Prostate cancer susceptibility locus on chromosome 1q: a confirmatory study.

Background: Recent recognition that a predisposition to prostate cancer can be inherited has led to a search for specific genes associated with the disease. Through a study of families with three or more affected first-degree relatives, a region on the long arm of chromosome 1 (i.e.

Biologic variability of prostate-specific antigen and its usefulness as a marker for prostate cancer: effects of finasteride. The Finasteride PSA Study Group.

Objectives: The effects of finasteride on prostate-specific antigen (PSA) variability and usefulness in prostate cancer detection were examined.

Methods: Percent change and crossover of PSA levels between the low (1.0 to 3.

The relationship between prostatic intraepithelial neoplasia and prostate cancer: critical issues.

Purpose: Prostatic intraepithelial neoplasia (PIN) is often considered to be a premalignant lesion and the main precursor of invasive carcinoma of the prostate. We evaluated the evidence for and against PIN as a premalignant lesion and determined guidelines for the clinical management of PIN.

Materials And Methods: Literature analysis of histopathological, morphometric, phenotypic and molecular genetic evidence of progression and of clinical findings regarding PIN was done.