Characterization of human blood dendritic cell subsets

Dendritic cells (DCs) are key antigen-presenting cells for stimulating immune responses and they are now being investigated in clinical settings. Although defined as lineage-negative (Lin(-)) HLA-DR(+) cells, significant heterogeneity in these preparations is apparent, particularly in regard to the inclusion or exclusion of CD14(+), CD16(+), and CD2(+) cells. This study used flow cytometry and a panel of monoclonal antibodies (mAbs), including reagents from the 7th Leukocyte Differentiation Antigen Workshop, to define the cellular composition of 2 standardized peripheral blood mononuclear cell (PBMCs)-derived Lin(-) HLA-DR(+) preparations. Lin(-) cells were prepared from PBMCs by depletion with CD3, CD14, CD19, CD11b, and either CD16 or CD56 mAbs. Analysis of the CD16-replete preparations divided the Lin(-) HLA-DR(+) population into 5 nonoverlapping subsets (mean +/- 1 SD): CD123 (mean = 18.3% +/- 9.7%), CD1b/c (18.6% +/- 7.6%), CD16 (49.6% +/- 8.5%), BDCA-3 (2.7% +/- 1.4%), and CD34 (5.0% +/- 2.4%). The 5 subsets had distinct phenotypes when compared with each other, monocytes, and monocyte-derived DCs (MoDCs). The CD85 family, C-type lectins, costimulatory molecules, and differentiation/activation molecules were also expressed differentially on the 5 Lin(-) HLA-DR(+) subsets, monocytes, and MoDCs. The poor viability of CD123(+) DCs in vitro was confirmed, but the CD16(+) CD11c(+) DC subset also survived poorly. Finally, the individual subsets used as stimulators in allogeneic mixed leukocyte reactions were ranked by their allostimulatory capacity as CD1b/c > CD16 > BDCA-3 > CD123 > CD34. These data provide an opportunity to standardize the DC populations used for future molecular, functional and possibly even therapeutic studies.     

Detection of circulating crosslinked fibrin derivatives by a heat extraction-SDS gradient gel electrophoretic technique

A technique has been developed to identify and quantitate unique plasmic degradation products of crosslinked fibrin in plasma. In this method, fibrin derivatives are extracted by heat precipitation and dissolved with disulfide bond reduction, after which the crosslinked gamma-gamma chain remnants are identified by SDS-polyacrylamide gradient gel electrophoresis and quantitated by densitometric analysis. A heterogenous group of gamma-gamma chains with molecular weights between 100,000 and 76,000 daltons was identified in lysates of crosslinked fibrin during plasmic degradation in vitro. Three stages of crosslinked fibrin degradation have been arbitrarily defined based primarily on the extent of degradation of these gamma-gamma polypeptide chains. As little as 20 microgram of crosslinked fibrin digests added to 1 ml of normal plasma could be detected by the heat-extraction--gel- electrophoresis technique, identifying the gamma-gamma derivatives with molecular weights of 96,000, 86,000, 82,000, and 76,000 daltons. Plasmic derivatives of gamma-gamma chains were not found in normal plasma, but they were identified in the plasma of patients with disseminated intravascular coagulation and deep-vein thrombosis, both before and in increased quantity during successful thrombolytic therapy.     

Interactions of ACTH and TGFβ on monocyte proliferation: Implications for trauma and burn patients

The purpose of this study was to try to elucidate a possible biobehavioral mechanism associated with decreased immune function in trauma patients by determining whether there is an interaction between the effects of ACTH, a stress hormone, and TGFβ, a cytokine, on peripheral blood lymphocyte proliferation. Peripheral mononuclear lymphocytes (PMLs) from healthy donors were preincubated with varying concentrations of ACTH for 24 hr, stimulated with Conconavalin A and increasing concentrations of TGFβ, and incubated for 72 hr. Proliferation was assayed by tritiated thymidine incorporation. A parallel aliquot of PMLs were incubated in the presence of ACTH to determine the direct effect of ACTH on mononuclear cell TGFβ production. While harvested supernatant from cells incubated in the presence of ACTH did not contain any detectable TGFβ, ACTH as well as TGFβ were found to significantly decrease cellular proliferation independent of one another. An even greater decrease in cellular proliferation was found when both ACTH and TGFβ were used, compared to either ACTH or TGFβ alone. These results suggest a biobehavioral interaction between ACTH and TGFβ at the cellular level and that interactions to relieve stress may assist in improving function and recovery from trauma. © 1996 John Wiley & Sons, Inc.     

OraSure InteliSwab™ Rapid Antigen Test Performance with the SARS-CoV-2 Variants of Concern—Alpha,Beta, Gamma,Delta, and Omicron

The emergence of SARS-CoV-2 in the human population and the resulting COVID-19 pandemic have led to the development of various diagnostic tests. The OraSure InteliSwab^™ COVID-19 Rapid Test is a recently developed and FDA emergency use-authorized rapid antigen-detecting test that functions as a lateral flow device targeting the nucleocapsid protein. Due to SARS-CoV-2 evolution, there is a need to evaluate the sensitivity of rapid antigen-detecting tests for new variants, especially variants of concern such as Omicron. In this study, the sensitivity of the OraSure InteliSwab^™ Test was investigated using cultured strains of the known variants of concern (VOCs, Alpha, Beta, Gamma, Delta, and Omicron) and the ancestral lineage (lineage A). Based on dilution series in cell culture medium, an approximate limit of detection for each variant was determined. The OraSure InteliSwab^™ Test showed an overall comparable performance using recombinant nucleocapsid protein and different cultured variants, with recorded limits of detection ranging between 3.77 × 10⁵ and 9.13 × 10⁵ RNA copies/mL. Finally, the sensitivity was evaluated using oropharyngeal swabs from Syrian golden hamsters inoculated with the six VOCs. Ultimately, the OraSure InteliSwab^™ COVID-19 Rapid Test showed no decrease in sensitivity between the ancestral SARS-CoV-2 strain and any VOCs including Omicron.     

Discussion of fertility preservation with newly diagnosed patients: oncologists’ views

Introduction Although physician discussion with patients regarding fertility preservation (FP) options prior to cancer treatment can provide important information for survivors concerning their future fertility, little is known about the extent to which physicians discuss FP with patients. This qualitative study sought to identify current physician FP communication practices and determine factors that may impact communication efforts regarding FP. Materials and methods Qualitative data were collected using semi structured interviews with 16 physicians practicing at a major cancer center in the South. Results All providers were board certified in medical oncology, radiation oncology or surgical oncology. The main factors that emerged from qualitative analysis included distinct variations in quality of discussion about FP, knowledge of FP resources, attitudes, practice behaviors and perceptions of patient characteristics. Discussion While most physicians discussed potential fertility loss as a side effect of cancer treatment, few provided information to patients about preserving fertility. Patient characteristics such as gender and cancer site may impact the discussion, as well as system factors such as costs of procedures and access to FP resources. Education and training for physicians about FP options for cancer patients, particularly females, may promote discussion of FP. In addition, system barriers related to availability and affordability of FP resources must also be addressed. Implications for cancer survivors Physicians should consider providing patients with timely, understandable information related to their FP options, prior to the administration of treatment. Such discussions may lead to improved quality of life for individuals as they transition from patients to survivors.     

Identification of the human P450 enzymes involved in the in vitro metabolism of the synthetic steroidal hormones Org 4060 and Org 30659

1. The type of human P450 enzymes involved in the in vitro metabolism of Org 4060 and Org 30659, two synthetic steroidal hormones currently under clinical development by NV Organon for use in oral contraceptive and hormone replacement therapy, was investigated. 2. Both steroids were mainly hydroxylated at the 6beta-position in incubations with human liver microsomes. 3. The results from experiments with supersomes, correlation studies as well as inhibition studies with ketoconazole, a selective inhibitor of CYP3A, strongly suggest that the CYP3A family plays a significant role in the 6beta-hydroxylation of both steroids. 4. Measurements of kinetic parameters of P450 enzymes that could metabolize both steroids, combined with the fact that CYP3A4 is known to be the most abundant P450 enzyme in the human liver, indicate that CYP3A4 will be of major importance for the in vivo human metabolism of Org 4060 and Org 30659.     

The bottom line: Outcomes after conservation treatment in anal cancer

At the Department of Radiation Oncology, Westmead Hospital, between 1980 and 2000, 60 patients with squamous cell carcinoma of anal canal or margin (including 15 with Stage IIIA or IIIB) were treated radically; 55 received chemoradiation (89% were prescribed mitomycin C and 5‐fluorouracil). Five‐year overall survival was 64% (95% confidence interval (CI): 48–79%), with a median survival of 9.75 years (median follow up 5.6 years, range 5 months to 22.5 years). Ten patients have died of disease. At 2 years the local control rate was 86%, and colostomy‐free survival was 83%. Relapse after 2 years was uncommon. Tumour size was the main factor driving outcomes, especially survival. Patients with larger tumours (T > 4 cm) had a hazard ratio for survival of 5.7 (95% CI: 1.8–17). Fourteen (24%) patients experienced treatment interruptions as a result of acute toxicity, including one death from neutropoenic sepsis. Seven (12%) patients, in total, experienced one or more late toxicities, grade 3 or above, including four women (all postmenopausal) who developed a radiation‐induced bone injury. Most patients with anal cancer can expect to retain a functional sphincter after chemoradiation/radiation. Further studies are in progress to determine the optimal chemoradiation protocol.