Doses Of Ultraviolet Radiation Research Articles

Acute UV Irradiation Increases Heparan Sulfate Proteoglycan Levels in Human Skin

Glycosaminoglycans are important structural components in the skin and exist as various proteoglycan forms, except hyaluronic acid. Heparan sulfate (HS), one of the glycosaminoglycans, is composed of repeated disaccharide units, which are glucuronic acids linked to an N-acetyl-glucosamine or its sulfated forms. To investigate acute ultraviolet (UV)-induced changes of HS and HS proteoglycans (HSPGs), changes in levels of HS and several HSPGs in male human buttock skin were examined by immunohistochemistry and real-time quantitative polymerase chain reaction (qPCR) after 2 minimal erythema doses (MED) of UV irradiation (each n = 4-7). HS staining revealed that 2 MED of UV irradiation increased its expression, and staining for perlecan, syndecan-1, syndecan-4, CD44v3, and CD44 showed that UV irradiation increased their protein levels. However, analysis by real-time qPCR showed that UV irradiation did not change mRNA levels of CD44 and agrin, and decreased perlecan and syndecan-4 mRNA levels, while increased syndecan-1 mRNA level. As HS-synthesizing or -degrading enzymes, exostosin-1 and heparanase mRNA levels were increased, but exostosin-2 was decreased by UV irradiation. UV-induced matrix metalloproteinase-1 expression was confirmed for proper experimental conditions. Acute UV irradiation increases HS and HSPG levels in human skin, but their increase may not be mediated through their transcriptional regulation.

Neonatal mice do not have increased sensitivity to induction of squamous cell carcinomas

Squamous cell carcinoma (SCC) is linked with the lifelong cumulative effect of ultraviolet radiation (UVR). In contrast, epidemiological data have shown that sunburn in childhood is a stronger risk factor for cutaneous malignant melanoma than continuous UVR, indicating a higher carcinogenic sensitivity early in life. We investigated how a high neonatal dose of UVR affects the development of SCC in mice irradiated later in life. We used simulated solar radiation (sun) and solarium radiation (solarium). Ninety-nine C3.Cg/TifBomTac-immunocompetent hairless mice received 0, 25 or 35 standard erythema doses (SED) UVR when they were 4 days old followed by 4 SED sun or 4 SED solarium three times/weekly from 9 weeks of age. Tumours developed faster in mice treated with 35 SED UVR + 4 SED sun compared with 4 SED sun, but no change was observed in the cumulative dose required to achieve tumours. Tumours also developed faster in mice treated with 35 SED UVR + 4 SED solarium compared with 4 SED solarium, and a difference was also observed in the cumulative dose required to achieve tumours. If the Skin Cancer Utrecht-Philadelphia-murine spectrum was used to weigh the delivered irradiance instead of the International Commission on Illumination erythema action spectrum, tumours developed after the same accumulated dose. In conclusion, this study does not indicate increased sensitivity to induction of SCC early in life.

Repeated small perturbation approach reveals transcriptomic steady states.

The study of biological systems dynamics requires elucidation of the transitions of steady states. A “small perturbation” approach can provide important information on the “steady state” of a biological system. In our experiments, small perturbations were generated by applying a series of repeating small doses of ultraviolet radiation to a human keratinocyte cell line, HaCaT. The biological response was assessed by monitoring the gene expression profiles using cDNA microarrays. Repeated small doses (10 J/m2) of ultraviolet B (UVB) exposure modulated the expression profiles of two groups of genes in opposite directions. The genes that were up-regulated have functions mainly associated with anti-proliferation/anti-mitogenesis/apoptosis, and the genes that were down-regulated were mainly related to proliferation/mitogenesis/anti-apoptosis. For both groups of genes, repetition of the small doses of UVB caused an immediate response followed by relaxation between successive small perturbations. This cyclic pattern was suppressed when large doses (233 or 582.5 J/m2) of UVB were applied. Our method and results contribute to a foundation for computational systems biology, which implicitly uses the concept of steady state.

Ground-based Observations of Ultraviolet and Total Solar Radiation in Shenyang, Northeast China

Objective This work explores the diurnal variation of Solar ultraviolet radiation (UVR) and total solar radiation (TSR) in northeast China, using daily observations of UVR and TSR in Shenyang. Methods UVR and TSR measurements were carried out from March 1st, 2006 to December 31st, 2009 in Shenyang, Liaoning province, China (41°51′ N, 123°27′ E). Results Both TSR and UVR showed seasonal variation, reaching the highest levels in summer and the lowest in winter. They showed the greatest fluctuation in summer and autumn. The irradiance of TSR and UVR on clear days around the equinoxes and solstices increased substantially compared with the mean seasonal irradiance, especially in autumn. The whole day accumulated dose of UVR in winter was far less than that during the middle part of a summer day (i.e. between 10:00 and 14:00). It was also less than the accumulated summer dose of morning and afternoon (i.e. between 8:00 and 10:00 and 14:00 and 16:00). Conclusion The instant irradiance and daily accumulated amount of UVR are low in Shenyang, especially in autumn and winter. Thus concern about the health effects arising because shortage of UVR in northeast China is warranted.

Effect of ultraviolet filters on skin superoxide dismutase activity in hairless mice after a single dose of ultraviolet radiation

Organic sunscreens may decrease their protective capability and also behave as photo-oxidants upon ultraviolet radiation (UVR) exposure. The present study investigated the effect of a cream gel formulation containing the UV filters benzophenone-3, octyl methoxycinnamate, and octyl salicylate on skin superoxide dismutase (SOD) after a single dose of UVR (2.87J/cm2). The retention of these UV filters was first evaluated in vivo using hairless mice to guarantee the presence of the filters in the skin layers at the moment of irradiation. The in vivo effect of the UV filters on skin SOD was then assayed spectrophotometrically via the reduction of cytochrome c. The cream gel formulation promoted the penetration of the three UV filters into the epidermis and the dermis at one hour post-application. A significant decrease in SOD activity was observed in irradiated animals treated with sunscreen formulation. However, no effect on SOD activity in skin was observed by the isolated presence of the sunscreens, the formulation components, or the exposure to UVR. The sunscreens may have formed degradation products under UVR that may have either inhibited the enzyme or generated reactive species in the skin.

Evolution of protein concentration in the rat lens after in vivo exposure to close‐to‐threshold dose ultraviolet radiation

AbstractPurpose To investigate the short term cataract development and protein concentration in the rat lens after in vivo close‐to‐threshold dose exposure to ultraviolet radiation (UVR) around 300 nm.Methods Three groups of 10 Sprague Dawley rats were unilaterally exposed to 8 kJ/m2 UVR‐300 nm for 15 minutes, and a forth group of 10 rats was kept without UVR exposure as non‐exposed control animals. The exposed animals were sacrificed at 1, 3 and 7 days after exposure. Both lenses from all animals were extracted and photographed and the intensity of forward light scattering was measured quantitatively. Whole lens was homogenized and protein concentration was determined spectrophotometrically.Results All exposed lenses developed cataract. Lens light scattering increased throughout 7 days after UVR exposure. The difference of protein concentration between exposed lens and contralateral non‐exposed lens was slightly decreased at 1 day after exposure and then gradually normalized back to baseline within 7 day after exposure.Conclusion The in vivo close‐to‐threshold dose UVR induced cataract and protein changes are not paralleled. The altered protein concentration is normalized within 7 days after ultraviolet irradiation.

The effects of grape seeds polyphenols on SKH-1 mice skin irradiated with multiple doses of UV-B

The study investigated the protective activity of red grape seeds ( Vitis vinifera L, Burgund Mare variety) ( BM) extracts in vivo on multiple doses of ultraviolet radiation (UV)-B-induced deleterious effects in SKH-1 mice skin. Eighty 8-weeks-old female SKH-1 mice were divided into 8 groups: control, vehicle, UV-B irradiated, vehicle + UV-B irradiated, BM 2.5 mg polyphenols (PF)/cm 2 + UV-B irradiated, BM 4 mg PF/cm 2 + UV-B irradiated, UV-B + BM 2.5 mg PF/cm 2, UV-B + BM 4 mg PF/cm 2. The extract was applied topically before or after each UV-B exposure (240 mJ/cm 2), for 10 days consecutively. The antioxidant activity of BM extract is higher than gallic acid ( k BM = 0.017, k gallic acid = 0.013). Multiple doses of UV-B generated the formation of cyclobutane pyrimidine dimers (CPDs) and sunburn cells, increased glutathione peroxidase (GPx) and catalase (CAT) activities respectively glutathione (GSH) and IL-1β levels in skin. In group treated with 2.5 mg PF/cm 2 before UV-B irradiation BM extract inhibited UV-B-induced sunburn cells, restored the superoxide dismutase (MnSOD) activity, increased insignificantly CAT and GPx activities and reduced IL-1β level. The BM 4.0 mg PF/cm 2 treatment decreased GSH level and reduced the percentage of CPDs positive cells in skin. Both doses of BM extract administered after UV-B irradiation increased the MnSOD and GPx activities and reduced the formation of sunburn cells in skin. Our results suggest that BM extract might be a potential chemo-preventive candidate in reducing the oxidative stress and apoptosis induced by multiple doses of UV-B in skin.

Outdoor sports and risk of ultraviolet radiation-related skin lesions in children: evaluation of risks and prevention

Excessive ultraviolet (UV) radiation exposure can cause skin cancers, skin photoageing and cataracts. Children are targeted by sun-protection campaigns because high sun exposure and sunburn in childhood increase the risk of melanoma in adulthood. Little information is available about UV radiation risk and exposure in children who take part in outdoor sports. To evaluate the risk of developing UV radiation-induced skin lesions run by children who practise outdoor sports, and UV radiation exposure and sun-protection measures during a soccer tournament. Firstly, we evaluated the relationship between melanocytic naevus - a skin lesion linked with exposure to UV radiation - and outdoor sports in 660 11-year-old children. Secondly, we used the occasion of a 1-day soccer tournament held in the spring to evaluate UV radiation-protective measures used by soccer players and the public. We also evaluated the UV radiation index and cloud cover during the tournament, and calculated the UV radiation dose and minimal erythema dose depending on skin phototype. The naevus count and acquired naevus count measured over the 2 years of the study were higher in the 344 children who practised outdoor sports. Sun-protective measures were insufficient for soccer players and the public. This study shows that outdoor sports increase the risk of developing UV radiation-induced skin lesions in childhood. During a 1-day soccer tournament held in the spring, children and their parents were inadequately protected against the sun. These results suggest that sun-protection campaigns should be aimed at children who practise popular outdoor sports.

Does the Eye Benefit From Wearing Ultraviolet-Blocking Contact Lenses?

The human eye is exposed to toxic ultraviolet radiation (UVR) from sunlight and artificial sources. The UVR-induced damage occurs in ocular tissues from the corneal surface to the retina. Although the cornea and crystalline lens provide inherent UVR protection, the anterior ocular surface and the limbus, which contains stem cells, receive toxic levels of UVR from relatively short solar exposures. Shading headwear and some UVR-blocking sunglasses are designed to reduce direct solar exposure but may not protect the eye from diffuse ambient and surface reflected light. If the squint mechanism is reduced because of the reduction of visible light, the ocular surface is then exposed to ambient and reflected UVR. In addition, laterally incident radiation that is focused across the cornea onto the limbus, a phenomenon known as peripheral light focusing, can increase the dose at the nasal limbus by a factor of 20. The UVR-blocking contact lenses that cover the limbus provide protection from all sources of ocularly incident UVR. Although directly relating solar UVR dose to ocular damage is epidemiologically challenging, irradiation of ocular cell cultures can estimate the toxic effects of UVR exposure. The use of UVR-blocking contact lenses greatly increases the time the wearer can be exposed to solar UVR before a toxic ocular dose is reached. There is a need for the development of a scientifically rigorous, clinically applicable ocular protection factor metric, based not only on the transmittance of eyewear but on the protection afforded from the total UVR field and the length of that exposure.

Prodigiosin from Vibrio sp. DSM 14379; A New UV-Protective Pigment

Pigments such as melanin, scytonemin and carotenoids protect microbial cells against the harmful effects of ultraviolet (UV) radiation. The role in UV protection has never been assigned to the prodigiosin pigment. In this work, we demonstrate that prodigiosin provides a significant level of protection against UV stress in Vibrio sp. DSM 14379. In the absence of pigment production, Vibrio sp. was significantly more susceptible to UV stress, and there was no difference in UV survival between the wild-type strain and non-pigmented mutant. The pigment's protective role was more important at higher doses of UV irradiation and correlated with pigment concentration in the cell. Pigmented cells survived high UV exposure (324J/m(2)) around 1,000-fold more successfully compared to the non-pigmented mutant cells. Resistance to UV stress was conferred to the non-pigmented mutant by addition of exogenous pigment extract to the growth medium. A level of UV protection equivalent to that exhibited by the wild-type strain was attained by the non-pigmented mutant once the prodigiosin concentration had reached comparable levels to those found in the wild-type strain. In co-culture experiments, prodigiosin acted as a UV screen, protecting both the wild-type and non-pigmented mutants. Our results suggest a new ecophysiological role for prodigiosin.

Seeing Versus Saving

The research presented in this paper describes the measurement of light and ultraviolet energy within a special collections facility, with the goal of evaluating whether levels recommended for museums and archival collections are being exceeded during research usage. An Elsec 764 hand-held light meter was used to record the light intensity falling on collection material held within and without V-shaped book mounts and with sequential lights turned on, as occurs in collections’ use. The authors developed a simple algebraic formula to calculate cumulative doses of light and incident ultraviolet radiation to determine how many hours collection material could be accessed and illuminated before damage could be expected. The authors calculated the maximum cumulative doses possible based on numbers of access hours and compared these to recommended doses for sensitive media as a monitoring strategy for the long-term preservation of light sensitive special collection materials. The results from this study suggest that the light levels evaluated are not in excess of recommended values and that the use of book mounts reduces the amount of light falling on collection material. Monitoring actions are recommended for institutions wishing to replicate the study.

Cutaneous Melanoma and Endogenous Hormonal Factors: A Large French Prospective Study

To assess the role of endogenous hormonal factors on melanoma, the authors conducted a prospective analysis of 91,972 French women, aged 40-65 years at inclusion into the Etude Epidémiologique auprès de femmes de l'Education Nationale (E3N) cohort. Between 1990 and 2005, 460 melanoma cases were ascertained. Relative risks and 95% confidence intervals were computed by using Cox proportional hazards regression models. Risks of melanoma were reduced in women with ≥ 15 years at menarche (relative risk (RR) = 0.67, 95% confidence interval (CI): 0.46, 0.97, compared with 13-14 years), irregular menstrual cycles (RR = 0.52, 95% CI: 0.31, 0.89, compared with regular cycles of 25-31 days), <48 years at natural menopause (RR = 0.70, 95% CI: 0.48, 1.02, compared with 48-51 years), and shorter ovulatory life (RR = 0.51, 95% CI: 0.28, 0.91, for <33 years compared with ≥ 39 years). Modest inverse associations were observed with parity, as well as number of pregnancies and miscarriages. There was no evidence of an association between melanoma risk and age at first birth or pregnancy, age at last birth, time since last birth, breastfeeding duration, age at menstruation regularity, or menopausal status. Results did not significantly differ according to ambient ultraviolet radiation dose and melanoma site or subtype. These findings from a large prospective cohort may suggest a reduced melanoma risk associated with decreased exposure to ovarian hormones.

Different Sensitivities of Whitefish (Coregonus Lavaretus) and Northern Pike (Esox Lucius) Eleutheroembryos to Photoinduced Toxicity of Polycyclic Aromatic Hydrocarbons

The photoinduced toxicity of two polycyclic aromatic hydrocarbons (PAHs), retene (RET) and pyrene (PYR), to the eleutheroembryos of whitefish and northern pike was studied. Fish were exposed to three concentrations of RET and PYR, and irradiated with ultraviolet radiation (UVR) or visible light for 3 h on two consecutive days. UVR covered the absorption maxima of RET and PYR at UVB and UVA, the daily UVR doses were 30 and 28 kJ m−2, respectively. After 72 h, mortality and behavioral abnormalities were observed. Survivors were analyzed for the proteins of CYP1A, CYP3A, Hsp70, and the activity of glutathione reductase (GR). While neither PAHs nor UVR on their own were lethal, simultaneous exposure caused mortality. The whitefish were more sensitive than the pike to the photoinduced toxicity, and PYR was much more phototoxic than RET. RET induced Hsp70 in both species, and PYR induced GR without UVR in pike.

The influence of low and moderate doses of ultraviolet irradiation produced by the UFA-1 device possessing the wavelength of 350 to 400 nm on the parameters of skin inflammation in patients suffering atopic dermatitis

The performed investigation has shown that there is an increase of the content of CD4+ lymphocytes and an increased expression of γ-interferon in the skin of patients with atopic dermatitis comparing to healthy volunteers. Therapy with the use of UFA-1 produced more marked levels of γ-interferon positive cells in the skin (р = 0.004) in patients with clinical recovery compared to subjects with an achieved improvement, as well as a substantial reduction of the Т-helpers level within the effected skin areas (р = 0.009).

The Vitamin D Debate: Translating Controlled Experiments into Reality for Human Sun Exposure Times

Exposure to sunlight, specifically the ultraviolet radiation, has both positive and negative health effects. Maximizing the benefits (vitamin D synthesis) while minimizing the damage is a multifaceted problem in which many of the elements are poorly quantified. Here we show how rigorously conducted large sample size laboratory studies of the effect of ultraviolet radiation dose on vitamin D status can be applied to real-life situations. This was achieved by modeling the radiation incident on different surfaces for different solar locations, and equating with the controlled exposures in the laboratory studies. Results from both model and experimental data show that relatively short exposures of a modest amount of unprotected skin to summer sunlight in northern climes, on a regular basis during lunchtime hours, increases vitamin D to sufficiency status (≥20 ng mL(-1) ) in the white Caucasian population. While both sun exposure conditions and human skin responses are variable in real life, these quantitative findings provide a guide for authorities devising sunlight exposure recommendations.

Highlights of This Issue

Chang et al., Page 23Two studies appearing in this issue evaluate whether cancer risk alleles that were originally identified in populations of European ancestry are also relevant in other patient populations. In one study, He and colleagues examined several colorectal cancer (CRC) risk variants in a multiethnic sample comprised of European American, African American, Native Hawaiian, Japanese American and Latino men and women. He and colleagues confirmed the associations of several CRC/adenoma risk alleles in European Americans and found statistically significant cumulative effects of risk alleles on CRC/adenoma risk in all populations except Japanese Americans. In another study, Chang and colleagues evaluated 27 GWAS-identified loci in prostate cancer patients of African descent. The authors found several significant associations in the African population and these associations were in the same direction and of similar magnitude as those reported in men of European descent. Although several prostate cancer risk alleles were confirmed in men of African descent, not all risk variants were validated. This may be due to different contributions of genetic and non-genetic factors in prostate cancer in men of African or European descent. Both of these studies stress the importance of evaluating the epidemiologic architecture of genetic risk factors in different populations.The medication metformin lowers elevated insulin levels among diabetic patients. Previous studies showed a protective effect of metformin on breast cancer risk. To expand on this work, Bosco and colleagues used Danish medical registries to conduct a nested case-control study evaluating the effect of metformin on breast cancer risk among peri- and postmenopausal type 2 diabetic women. They found that metformin users were indeed less likely to be diagnosed with breast cancer than non-metformin users. These data support the hypothesis that metformin may be a candidate breast cancer prevention agent.Little is known about the relationship between pre- and postmenopausal breast cancer and the combined intake of vitamin D from diet, supplements and sun exposure. To address this, Engel and colleagues evaluated the association between vitamin D intake and mean daily Ultraviolet Radiation dose (UVRd) with the risk of breast cancer among French women. They report that dietary and supplemental vitamin D intakes were not associated with breast cancer risk. Postmenopausal women with high vitamin D intake and living in regions of France with high UVRd, however, had significantly lower breast cancer risk. This study suggests that there may be a Vitamin D threshold exposure for breast cancer prevention.

Joint Effects of Dietary Vitamin D and Sun Exposure on Breast Cancer Risk: Results from the French E3N Cohort

Ecological studies have suggested that vitamin D production through ultraviolet (UV) solar irradiance could reduce breast cancer (BC) risk. Although studies restricted to dietary vitamin D intake have provided inconsistent results, little is known about the relationship between pre- and postmenopausal BC and combined intakes from diet, supplements, and sun exposure. Cox proportional hazards regression models evaluated the association between vitamin D intakes, mean daily ultraviolet radiation dose (UVRd) at the place of residence and risk of BC among 67,721 women of the French E3N cohort. All analyses were stratified on menopausal status taking into account important confounders including calcium consumption. During 10 years of follow-up, a total of 2,871 BC cases were diagnosed. Dietary and supplemental vitamin D intakes were not associated with BC risk; however, in regions with the highest UVRd, postmenopausal women with high dietary or supplemental vitamin D intake had a significantly lower BC risk as compared with women with the lowest vitamin D intake (HR = 0.68, 95% CI: 0.54-0.85, and HR = 0.57, 95% CI: 0.36-0.90, respectively). Our results suggest that a threshold of vitamin D exposure from both sun and diet is required to prevent BC and this threshold is particularly difficult to reach in postmenopausal women at northern latitudes where quality of sunlight is too poor for adequate vitamin D production. Prospective studies should further investigate associations between BC risk, vitamin D status and sunlight exposure.

Analysis of transmitted optical spectrum enabling accelerated testing of multijunction concentrating photovoltaic designs

Concentrating photovoltaic (CPV) technology has recently gained interest based on its scalability and expected low levelized cost of electricity. The reliability of encapsulation materials used in CPV systems, however, is not well established. For example, the present qualification test for CPV modules includes only real-time ultraviolet (UV) exposure, i.e., methods for accelerated UV testing have not yet been developed. To better define the stress inherent to CPV systems, the UV and infrared spectra transmitted through representative optical systems were evaluated. Measurements of optical components are used to assess expected optical performance and quantify damaging optical exposure. Optical properties (transmittance, refractive index, reflectance, and absorptance) of candidate materials (including PMMA, soda-lime glass, borosilicate glass, and quartz refractors), components (including Ag- and Al-enabled reflectors), and encapsulants (including EVA, ionomer, PDMS, PPMS, polyolefin, and PVB) were identified. The activation spectrum was calculated for the representative optical systems using an assumed action spectrum to compare the expected damaging dose of UV radiation delivered to the cell encapsulation. The dose and flux analysis identifies the significance of IR relative to UV exposure for CPV systems. Because UV light is typically more highly attenuated, the UV dose within the encapsulation may not greatly exceed the unconcentrated global solar condition, but the thermal load scales nearly directly with the geometric concentration. Relative to a previous analysis for crystalline silicon cell technology, the analysis here is performed for III-V multijunction technology. Novel aspects here also include additional materials (such as TPU encapsulation) and additional components (transmission through silicone on glass lenses, antireflective coatings, and the front glass used with reflective systems, as well as reflection off of the cell).

Effects of sunscreen on human skin’s ultraviolet radiation tolerance

To observe the alteration ultraviolet radiation (UVR) of skin's tolerance after its exposure to the small dose of UVR under the protection of sunscreen. Eleven subjects who applied sunscreen were exposed to 0.75 dose minimal persistent pigment darkening (MPPD) and minimal erythema dose (MED) by the Phototherapy Unit for 4 weeks. Each week their MPPDs and MEDs were measured by solar simulator. Meanwhile, SPECTCOLOMETER® and VISIOSCAN VC98® were used to detect the test areas and control areas. The values of MPPD and MED increased significantly after the exposure to UVR. But there were no visible changes on the surface of skin's texture. With the protection of sunscreen, the UVR tolerance of skin was greatly increased after the skin's exposure to the small dose UV.

Ultraviolet Radiation Dose to be Applied in Recipient Zebrafish Embryos for Germ‐line Chimaerism is Strain Dependent

Germ-line chimaerism is a powerful technique that has proved to be useful to produce viable gametes when transplanted blastomeres colonize the germinal ridges in recipient embryos and obtaining offspring from such transplanted cells. In fish, ionizing radiations were commonly used for embryo penalization to cancelling the cell participation of recipient embryos in development and in gamete production. The ultraviolet (UV) radiation when compared with other radiation types is cheaper, easier and no special installations are required for its use. So, the aim of this work was to establish the optimal UV radiation dose to be applied in zebrafish embryos at mid-blastula transition stage of development, in order to use them as penalized recipient embryos in futures chimaerism assays. A UV germicide lamp was used as radiation source (0.529 mW/cm(2)). Four exposure levels and three exposure times of UV radiation were tested. The survival rates obtained with the non-dechorionated embryos without lid group suggested that it could be the optimal exposure level to achieve the objective proposed. With the obtained results, we concluded that this UV radiation dose for 60 and 30 s are optimal parameters to penalize recipient wild and gold strain zebrafish embryos, respectively in chimaerism assays, but without involving their survival and apparently normal development.