A Needs Assessment Among Transgender Patients at an LGBTQ Service Organization in Texas.

Purpose: Transgender adults have difficulty accessing health care due to multiple barriers. This study examined the health care-related needs of transgender patients in Dallas, Texas. Methods: This study examined cross-sectional data from a survey completed by 62 patients who identified as transgender. Results: Many participants reported depression (50%) and anxiety (51%). Over half did not receive preventive screenings (60%) or health care (61%) elsewhere. One-third of patients felt their primary care physician outside the clinic was not transgender-friendly. Conclusion: These findings provide evidence that transgender patients demonstrate increased reported mental health disorders and decreased access to medical care.

Comparison of time to PRRSv-stability and production losses between two exposure programs to control PRRSv in sow herds.

To control and eliminate porcine reproductive and respiratory syndrome virus (PRRSv) from breeding herds, some veterinarians adopt a strategy called load-close-expose which consists of interrupting replacement pig introduction for several months and exposing the pigs to a replicating PRRSv. This was a prospective quasi-experiment that followed 61 breeding herds acutely infected with PRRSv that adopted one of two exposure programs: modified-live virus (MLV) vaccine or live-resident virus inoculation (LVI). Treatment groups (load-close-expose with MLV or LVI) were compared for: (a) time-to-PRRSv stability (TTS), defined as time in weeks it took to produce PRRSv negative pigs at weaning; (b) the time-to-baseline production (TTBP), defined using statistical process control methods to represent time to recover to the number of pigs weaned per week that herds had prior to PRRSv-detection; and (c) the total production loss in terms of number of pigs weaned per week. TTS and TTBP were compared between treatments using survival analysis. Day 1 of the program was considered to be the day that treatment was administered. Sampling at herds consisted of bleeding 30 due-to-wean piglets on a monthly basis. Serum was tested for PRRSv RNA by RT-PCR. Herds in which PRRSv was not detected over a 90-day period were classified as reaching stability. Multivariate analysis using proportional hazards regression was performed adjusting the effect of treatment on TTBP and TTS to 'severity of PRRSv infection', 'number of whole-herd exposures', 'days from PRRSv-detection to intervention', 'prior PRRSv-infection status' and 'veterinary clinic associated with the herd'. Total loss was compared between groups using multivariate regression analysis adjusted by selected covariates. The median TTS among participating herds was 26.6 weeks (25th to 75th percentile, 21.6-33.0 weeks). The overall TTBP was 16.5 weeks (range 0-29 weeks). The magnitude of production losses following whole-herd exposure averaged 2217 pigs not weaned/1000 sows and was correlated with TTBP. Herds in the MLV group recovered production sooner and had less total loss than herds in the LVI group. TTBP and TTS were significantly shorter and the total loss was significantly less in herds assisted by a specific veterinary clinic and herds that were infected with PRRSv in the 3 years prior to the study. This study provided new metrics to assist veterinarians to decide between methods of exposure to control and eliminate PRRSv from breeding herds.

Estimation of neutron-equivalent dose in organs of patients undergoing radiotherapy by the use of a novel online digital detector.

Neutron peripheral contamination in patients undergoing high-energy photon radiotherapy is considered as a risk factor for secondary cancer induction. Organ-specific neutron-equivalent dose estimation is therefore essential for a reasonable assessment of these associated risks. This work aimed to develop a method to estimate neutron-equivalent doses in multiple organs of radiotherapy patients. The method involved the convolution, at 16 reference points in an anthropomorphic phantom, of the normalized Monte Carlo neutron fluence energy spectra with the kerma and energy-dependent radiation weighting factor. This was then scaled with the total neutron fluence measured with passive detectors, at the same reference points, in order to obtain the equivalent doses in organs. The latter were correlated with the readings of a neutron digital detector located inside the treatment room during phantom irradiation. This digital detector, designed and developed by our group, integrates the thermal neutron fluence. The correlation model, applied to the digital detector readings during patient irradiation, enables the online estimation of neutron-equivalent doses in organs. The model takes into account the specific irradiation site, the field parameters (energy, field size, angle incidence, etc) and the installation (linac and bunker geometry). This method, which is suitable for routine clinical use, will help to systematically generate the dosimetric data essential for the improvement of current risk-estimation models.

Transient protection by peripheral benzodiazepine receptors during the early events of ultraviolet light-induced apoptosis.

The peripheral benzodiazepine receptor (PBR) is a mitochondrial protein involved in the formation of mitochondrial permeability transition (PT) pores which play a critical role during the early events of apoptosis. PBRs are located in many tissues and are strongly expressed in the superficial layers of human epidermis. PBRs play a protective role against free radical damage and PBR ligands modulate apoptosis. To investigate the role of PBR during the early events of ultraviolet (UV)-mediated apoptosis we compared the effects of UVB on PBR-transfected Jurkat cells and their wild type counterparts devoid of any PBR expression. Results indicate that early after UVB exposure (up to 4 h), PBR-transfected cells were more resistant to apoptosis and exhibited a delayed mitochondrial transmembrane potential drop, a diminished superoxide anions production, and a reduced caspase-3 activation. Taken together these findings suggest that PBR may regulate early death signals leading to UV induced apoptosis.

Ultraviolet AI exposure of human skin results in Langerhans cell depletion and reduction of epidermal antigen-presenting cell function: partial protection by a broad-spectrum sunscreen.

BACKGROUND: Ultraviolet (UV) B-induced effects on the skin immune system have been extensively investigated, but little is known regarding the immunological changes induced by UVA exposure of human skin. Recent data assessing the protection afforded by sunscreens against photoimmunosuppression stress the need for broad-spectrum sunscreens with an adequate UVA protection. OBJECTIVES: The purpose of this study was first to determine the changes observed in epidermal Langerhans cells (ELC) density and epidermal antigen-presenting cell (APC) activity after exposure of human skin to UVAI (340-400 nm) radiation, and secondly to assess the immune protection afforded in vivo by a sunscreen formulation containing a long wavelength UVA filter with a low UVA protection factor (UVA-PF = 3). METHODS: Epidermal cell (EC) suspensions were prepared from skin biopsies 3 days after exposure to a single dose of UVAI (either 30 or 60 J cm(-2)). RESULTS: Flow-cytometric analysis of EC suspensions revealed that exposure to 60 J cm(-2) UVAI resulted in a decreased number of ELC without infiltration of CD36+ DR+ CD1a- antigen-presenting macrophages into the epidermis, and a significant reduction of HLA-DR expression on viable ELC. In vivo exposure to both 30 and 60 J cm(-2) resulted in a decreased allogeneic CD4+ T-cell proliferation induced by UVAI-irradiated ECs. The sunscreen application partially prevented (57 +/- 9%) the decrease in epidermal allogeneic APC activity induced by 60 J cm(-2) UVAI. CONCLUSIONS: In vivo UVAI exposure of human skin results in a decreased number of ELC and in a downregulation of epidermal APC activity. This last effect is partially prevented by prior application of a sunscreen with a low UVAI-PF value. These results indicate that increasing the absorption of UV filters for long UVA wavelengths may lead to an improved immune protection.

Fluoropyrimidine sensitivity of human MCF-7 breast cancer cells stably transfected with human uridine phosphorylase.

The relationship between uridine phosphorylase (UP) expression level in cancer cells and the tumour sensitivity to fluoropyrimidines is unclear. In this study, we found that UP overexpression by gene transfer, and the subsequent efficient metabolic activation of 5-fluorouracil (5-FU) by the ribonucleotide pathway, does not increase the fluoropyrimidine sensitivity of MCF-7 human cancer cells.

Combination of thymidine phosphorylase gene transfer and deoxyinosine treatment greatly enhances 5-fluorouracil antitumor activity in vitro and in vivo.

We reported previously that 5-fluorouracil (FUra) efficacy could be enhanced by increasing tumoral thymidine phosphorylase (TP) activity. Potentiated TP yield was achieved by either transfecting cells with human TP gene (A. Evrard et al., Br. J. Cancer, 80: 1726-1733, 1999) or associating FUra with 2'-deoxyinosine (d-Ino), a modulator providing the tumors with TP cofactor deoxyribose 1-phosphate (J. Ciccolini et al., Clin. Cancer Res., 6: 1529-1535, 2000). The purpose of the present work was to study the effects of a combined modulation (TP gene transfer + use of d-Ino) on the sensitivity to FUra of the LS174T human colorectal cell line. Results showed a near 4000 times increase of cell sensitivity in vitro after double (genetic + biochemical) modulation. This potentiation of tumor response was accompanied by a total change in the FUra anabolic pathway with a 5000% increase of cytosolic fluorodeoxyuridine monophosphate, a stronger and longer inhibition of thymidylate synthase, and 300% augmentation of DNA damage. Besides, whereas thymidine failed to inhibit FUra cytotoxicity in LS174T wild-type cells, the potentiation of the antitumor activity observed in the modulating regimen was partly reversed by thymidine, indicative of thymidylate synthase as the main drug target. The impact of this double modulation was next investigated in xenograft-bearing nude mice. Results showed that whereas FUra alone was completely ineffective on wild-type tumor growth, the size of TP-transfected tumors in animals treated with the FUra/d-Ino combination was reduced by 80% (P < 0.05). Our results suggest that FUra exhibits stronger antiproliferative activity when activated via TP through the DNA pathway and that high tumoral TP activity therefore leads to enhanced sensitivity to fluoropyrimidines.

In vitro detection of indirect-acting genotoxins in the comet assay using Hep G2 cells.

The induction of DNA damage by four known promutagens (cyclophosphamide (CP), benzo(a)pyrene (BP), dimethylbenz(a)anthracene and 2-acetylaminofluorene (2AAF) was investigated on Hep G2 using the alkaline single cell electroporesis (SCGE) test, most often referred as the "comet assay". After a 3-day incubation, lysed cells embedded in agarose were electrophoresed under alkaline conditions, dyed with a SYBRgold fluorogen and analysed by the Komet software. Among the comet parameters provided by the image analysis program, statistical analysis did not identify any in particular that could best represent the DNA damages. All promutagens, when compared with the control, caused a statistically significant increase in DNA migration as determined by different parameters such as Olive tail moment, tail extent moment, tail/head or tail length. The data demonstrated the ability and the sensitivity of the comet assay when performed on Hep G2 in the detection of DNA damage induced by promutagens, and its suitability in mutagenicity testing in in vitro short-term assays.

[Clomipramine in the treatment of catatonia]. / Tratamiento de la catatonía con clomipramina.

The use of intravenous clomipramine gave rise to the remission of catatonia on the third or fourth day of treatment in two patients diagnosed as suffering from affective disorders. Therapeutic effect was maintained on switching to oral clomipramine. Clomipramine could be a therapeutic alternative in the management of catatonia associated with affective disorders.

Increased cytotoxicity and bystander effect of 5-fluorouracil and 5-deoxy-5-fluorouridine in human colorectal cancer cells transfected with thymidine phosphorylase.

5-Fluorouracil (5-FU) and 5'-deoxy-5-fluorouridine (5'-DFUR), a prodrug of 5-FU, are anticancer agents activated by thymidine phosphorylase (TP). Transfecting the human TP cDNA into cancer cells in order to sensitize them to these pyrimidine antimetabolites may be an important approach in human cancer gene therapy research. In this study, an expression vector containing the human TP cDNA (pcTP5) was transfected into LS174T human colon carcinoma cells. Eight stable transfectants were randomly selected and analysed. The cytotoxic effects of 5-FU and 5'-DFUR were higher in TP-transfected cells as compared to wild-type cells. The maximal decreases in the IC50 were 80-fold for 5-FU and 40-fold for 5'-DFUR. The increase in sensitivity to these pyrimidines of TP-transfected cells significantly correlated with the increase in both TP activity and TP expression. Transfected clone LS174T-c2 but not wild-type cells exhibited formation of [3H]FdUMP from [3H]5-FU. In addition the LS174T-c2 clone enhanced the cytotoxic effect of 5'-DFUR, but also that of 5-FU, towards co-cultured parental cells. For both anti-cancer agents, this bystander effect did not require cell-cell contact. These results show that both 5-FU or 5'-DFUR could be used together with a TP-suicide vector in cancer gene therapy.

The expression of peripheral benzodiazepine receptors in human skin: the relationship with epidermal cell differentiation.

The peripheral benzodiazepine receptor (PBR) is a protein of mitochondrial outer membranes utilizing porphyrins as endogenous ligands. PBR is part of a heteromeric receptor complex involved in the formation of mitochondrial permeability transition pores and in the early events of apoptosis. PBR may function as an oxygen-dependent signal generator; recent data indicate that these receptors may preserve the mitochondria of haematopoietic cell lines from damage caused by oxygen radicals. To identify PBRs in human skin, we used a specific monoclonal antibody directed against the C-terminus fragment of the human receptor. PBR immunoreactivity was found in keratinocytes, Langerhans cells, hair follicles and dermal vascular endothelial cells. Interestingly, confocal microscopic examination of skin sections revealed that PBR expression was strongly upregulated in the superficial differentiated layers of the epidermis. Ultrastructurally, PBRs were distributed throughout the cytoplasm but were selectively expressed on the mitochondrial membranes of epidermal cells. The elevated level of PBRs in the spinous layer was not associated with an increased number of mitochondria nor with an increased amount of mRNA as assessed by in situ hybridization on microautoradiographed skin sections. The present work provides, for the first time, evidence of PBR immunoreactivity in human skin. This mitochondrial receptor may modulate apoptosis in the epidermis; its increased expression in differentiated epidermal layers may represent a novel mechanism of natural skin protection against free radical damage generated by ultraviolet exposure.

Enhancement of 5-fluorouracil cytotoxicity by human thymidine-phosphorylase expression in cancer cells: in vitro and in vivo study.

Transferring a gene into cancer cells in order to sensitize them to drugs is an important approach in human cancer gene-therapy research. Thymidine phosphorylase (TP) is the first enzyme in the metabolic activation pathway of 5-fluorouracil (5-FU) to fluorodeoxyribonucleotides, thus, it could be used to increase the sensitivity of cancer cells to this anti-pyrimidine agent. In this study, an expression vector containing the human TP cDNA was transfected into C26 murine colon-carcinoma cells. Stable transfectants were selected; all showed increased TP activity, ranging from 2- to 10-fold when compared with wild-type cells. The in vitro sensitivity of transfectants to 5-FU and 5'-deoxy-5-fluorouridine (5'-DFUR) was enhanced, in agreement with the observed increase in TP activity. Then, tumors were generated by s.c. injection of TP-transfected or wild-type C26 cells in syngeneic BALB/c mice. 5-FU (25 mg/kg, i.p.) induced a growth delay of TP-transfected C26 tumors as compared with C26 wild-type tumors. These data suggest that TP could be transfected in tumor cells to increase the sensitivity to 5-FU for subsequent cancer gene therapy.

Immunosuppression induced by acute solar-simulated ultraviolet exposure in humans: prevention by a sunscreen with a sun protection factor of 15 and high UVA protection.

BACKGROUND: Cutaneous exposure to UVB radiation impairs the induction of contact hypersensitivity (CHS). Variable results have been found among studies examining the use of sunscreens to prevent UV-induced immunosuppression. OBJECTIVE: Our purpose was to determine whether solar-simulated exposure of human skin resulted in an impairment of CHS responses and whether the preapplication of an intermediate sun protection factor (SPF) sunscreen could prevent this locally UV-induced immunosuppression. METHODS: Irritant and CHS responses to dinitrochlorobenzene (DNCB) were randomly assessed in 160 human volunteers with or without UV exposure and with or without prior application of an SPF 15 sunscreen with high UVA protection. DNCB sensitization was performed 3 days after acute UV irradiation corresponding to 3 minimal erythema doses. RESULTS: After solar-simulated UV exposure, the percentage of positive responses to DNCB sensitization dropped from 95% to 50% (p = 0.003). Prior application of the sunscreen formulation did not modify the percentage of positive responses (90%) and maintained the immunization rate at 85% among volunteers exposed to UV. CONCLUSION: A localized sunburn can impair the afferent arm of CHS reactions in humans. The use of intermediate SPF sunscreens with high UVA protection adequately protects from the suppression of CHS responses that occurs after acute solar-simulated UV exposure.

Quantification of CD1a, HLA-DR, and HLA class I expression on viable human Langerhans cells and keratinocytes.

In order to determine precisely the cellular density of surface molecules that are critical for antigen presentation in human epidermis, we utilized a quantitative immunofluorescence indirect assay and performed flow cytometric analysis of human epidermal cell (EC) suspensions. We first demonstrated that Tricolor-labeled streptavidin coupled to Cy-5 (SA-TC) was a reliable marker for non viable EC and that SA-TC+ EC accounted for the frequent nonspecific background of fluorescence due to isotype controls binding, although Langerhans cells (LC) and Keratinocytes (Kc) express Fc receptors for IgG on their surfaces. These results indicate that quantification of cell surface antigens on human EC requires the concomitant use of a marker of viability. Multicolor flow cytometric analysis allowed us to quantify CD1 molecules and major histocompatibility complex (MHC) antigens on viable human LC and Kc. Our results demonstrated a weak expression of MHC class I molecules on viable LC (163 +/- 19 x 10(3) molecules/cell) compared to viable Kc (785 +/- 110 x 10(3) molecules/cell). Mean antigen density of HLA-DR and CD1a molecules on viable LC were 579 +/- 82 x 10(3) molecules/cell and 1600 +/- 133 x 10(3) molecules/cell, respectively. Quantitative flow cytometry of viable EC may be proposed to evaluate the number of membrane antigens whose level of expression is related to cellular maturation or activation that occurs in skin diseases.

An in vitro nucleoside analog screening method for cancer gene therapy.

Suicide genes that sensitize cells to drugs that are normally nontoxic at therapeutic levels represent an important approach in human gene therapy research. We have developed an in vitro screening assay to assess the modulation of nucleoside analogs after transfection of a vector expressing the herpes simplex virus thymidine kinase gene (HSV-TK). The thymidine kinase gene enhances nucleoside phosphorylation to nucleotides that kill cells by blocking DNA elongation. Cells lines used are 3T3-NIH fibroblasts (parental cells) and 3T3-TKc3 (HSV-TK gene-transfected 3T3-NIH). Two types of analysis are performed: a cytotoxicity assay, the neutral red uptake assay to assess the IC50 on the two cell lines, and an HPLC analysis coupled to a radiochemical flow detector to evaluate metabolic profiles after incubation of cells with tritiated analogs. Results show that cells expressing the HSV-TK gene are more sensitive than the parent cells to the effect of acyclovir or ganciclovir, the reference purine analog drugs, and also to the effect of pyrimidine analogs, bromodeoxyuridine, bromovinyldeoxyuridine, and ethyldeoxyuridine. Promising nucleoside analogs for gene therapy that can be achieved by HSV-TK could be evaluated using this model.

Comparison of three in vitro cytotoxicity assays for estimating surfactant ocular irritation.

Three in vitro cytotoxicity assays [neutral red uptake assay (NRU), MTT test and total protein content determination (TPC)] were analysed to assess their value for predicting the ocular irritancy potential of 20 surfactants. For each test, three established cell lines (SIRC rabbit corneal cells, Balb/c 3T3 and L929 mouse fibroblasts) were used. The concentration that induced 50% inhibition relative to controls (IC(50)) was calculated for each test, cell line and chemical. In vivo ocular irritancy data were compared with in vitro results. None of these assays provided a marked correlation of surfactant ocular irritation (Spearman or Pearson correlation coefficient lower than 0.65, P < 0.01 and moderate agreement with Kappa test). An IC(50) threshold of 700 mug/ml was set to discriminate between surfactant irritation and non-irritation. Three compounds were detected as false negatives (CHAPS, CHAPSO and sodium taurocholate), and two as false positives (Triton X155 and Brij 35).

Role of the French drug licensing authority in the prevention of influenza pandemics.

The French Drug Agency is responsible for the control and delivery of batch release certificates. In the case of an influenza pandemic, the use of inactivated vaccines, produced according to well-established procedures and controlled according to the European Pharmacopea and FDA requirements, will be strictly dependent on the necessary delays for production and controls. Mutual recognition between the National Control Laboratories in Europe might help in shortening the delays. If new, inactivated vaccines are produced either on cell cultures or by using genetically modified organisms, and if live attenuated vaccines are needed, it would be suitable to organize ad hoc working groups and international collaborative studies in fields of both research and regulation.