Edgar Frenk

Candida colonization and subsequent infections in critically III surgical patients

ObjectiveThe authors determined the role of Candida colonization in the development of subsequent infection in critically ill patients.DesignA 6-month prospective cohort study was given to patients admitted to the surgical and neonatal intensive care units in a 1600-bed university medical center. MethodsPatients having predetermined criteria for significant Candida colonization revealed by routine microbiologic surveillance cultures at different body sites were eligible for the study. Risk factors for Candida infection were recorded. A Candida colonization index was determined daily as the ratio of the number of distinct body sites (dbs) colonized with identical strains over the total number of dbs tested; a mean of 5.3 dbs per patient was obtained. All isolates (n = 322) sequentially recovered were characterized by genotyping using contour-clamped homogeneous electrical field gel electrophoresis that allowed strain delineation among Candida species. ResultsTwenty-nine patients met the criteria for inclusion; all were at high risk for Candida infection; 11 patients (38%) developed severe infections (8 candidemia); the remaining 18 patients were heavily colonized, but never required intravenous antifungal therapy. Among the potential risk factors for candidal infection, three discriminated the colonized from the infected patients—i.e., length of previous antibiotic therapy (p < 0.02), severity of illness assessed by APACHE II score (p < 0.01), and the intensity of Candida spp colonization (p < 0.01). By logistic regression analysis, the latter two were the independent factors that predicted subsequent candidal infection. Candida colonization always preceded infection with genotypically identical Candida spp strain. The proposed colonization indexes reached threshold values a mean of 6 days before Candida infection and demonstrated high positive predictive values (66 to 100%). ConclusionsThe intensity of Candida colonization assessed by systematic screening helps predicting subsequent infections with identical strains in critically ill patients. Accurately identifying high-risk patients with Candida colonization offers opportunity for intervention strategies.

Mutations of keratinocyte transglutaminase in lamellar ichthyosis

Lamellar ichthyosis is a severe congenital skin disorder characterized by generalized large scales and variable redness. Affected individuals in three families exhibited drastically reduced keratinocyte transglutaminase (TGK) activity. In two of these families, expression of TGK transcripts was diminished or abnormal and no TGK protein was detected. Homozygous or compound heterozygous mutations of the TGK gene were identified in all families. These data suggest that defects in TGK cause lamellar ichthyosis and that intact cross-linkage of cornified cell envelopes is required for epidermal tissue homeostasis.

Mutations in the rod domains of keratins 1 and 10 in epidermolytic hyperkeratosis.

Epidermolytic hyperkeratosis is a hereditary skin disorder characterized by blistering and a marked thickening of the stratum corneum. In one family, affected individuals exhibited a mutation in the highly conserved carboxyl terminal of the rod domain of keratin 1. In two other families, affected individuals had mutations in the highly conserved amino terminal of the rod domain of keratin 10. Structural analysis of these mutations predicts that heterodimer formation would be unaffected, although filament assembly and elongation would be severely compromised. These data imply that an intact keratin intermediate filament network is required for the maintenance of both cellular and tissue integrity.

Two genes contribute to different extents to the heme oxygenase enzyme activity measured in cultured human skin fibroblasts and keratinocytes: implications for protection against oxidant stress.

Abstract— Activation of expression of the heme oxygenase (HO) gene appears to be involved in a cellular defense system in mammalian cells. We now demonstrate that while HO‐1 mRNA levels are strongly inducible in dermal fibroblasts they are barely inducible in human epidermal keratinocytes following oxidative stress (UVA radiation and hydrogen peroxide). Paralleling this result was the observation that HO‐2 mRNA levels were low in dermal fibroblasts but were high in epidermal keratinocytes. In neither case was the HO‐2 gene inducible. The expression of the two HO genes led to enzymatic activity in both types of skin cells with an approximately 2.5‐fold higher level of enzymatic activity present in keratinocytes compared with fibroblasts derived from the same biopsy. In addition, ferritin levels, which have been found to be augmented via the HO‐dependent release of iron from endogenous heme sources, were two‐ to three‐fold higher in keratinocytes compared with matching fibroblasts. This higher ferritin pool would result in an enhancement of cellular iron sequestering capacity that may confer increased resistance to oxidative stress. Indeed, keratinocytes showed less UVA radiation‐dependent cell membrane damage than fibroblasts. These results are consistent with the hypothesis that HO expression in human epidermis and dermis is related to cellular defense mechanisms that operate in human skin.

Sunlight and carcinogenesis: Expression of p53 and pyrimidine dimers in human skin following UVA I, UVA I + II and solar simulating radiations

DNA damage by UV radiation plays an essential role in skin cancer induction. We report that even sub‐erythemal doses of solar simulating radiation, are capable of inducing substantial nuclear damage, namely pyrimidine dimers and p53 induction in human skin in situ<0R>. The quantity and distribution of p53 induced in human skin by UV radiation depended highly on the waveband and dose of UV used. Solar simulating radiation induced very high levels of p53 throughout all layers in epidermal keratinocytes 24 hr following an erythemal dose (230 ± 15.9/1000 cells), and the induction followed a dose response. Following UVA I + II and UVA I radiations, p53 expression was approximately half of that seen with equivalent biological doses of solar simulating radiation (63.5 ± 28.5 and 103 ± 15.9, respectively). Expression of p53 was seen in basal cell keratinocytes at lower doses of UVA, but all layers of the epidermis were affected at higher doses. Pyrimidine dimer induction, however, was seen to be the same for equivalent biological doses of UVA I, UVA I + II and solar simulating radiations, which coincides with previous findings that pyrimidine dimers initiate the erythemal response and are implicated in skin carcinogenesis. When equivalent biological doses of pure UVA are used with no UVB contamination, significant nuclear alterations occur in human skin in situ,<0R> which can approach those seen with UVB radiation. Our results suggest that DNA damage assessed in vivo<0R> by immunohistochemistry could provide a very sensitive endpoint for determining the efficacy of protective measures, such as sunscreens or protective clothing, against both UVB‐ and UVA‐induced damage in human skin. Int. J. Cancer 76:201–206, 1998.© 1998 Wiley‐Liss, Inc.

Screening for Antifungal Activity of Panamanian Plants

Crude extracts of Panamian plants (153 representing 28 species from 21 families) have been screened for antifungal activity against Candida albicans and Cladosporium cucumerinum. Activity was assessed in a semiquantitative fashion by bioautography on TLC plates. When tested at 100 pg, 15% of the extracts showed activity against one of the fungi, and 9% of the extracts were active against both test organisms. The most promising extracts originate from plants used in traditional medicine, such as Eursera simaruba (Burseraceae), Gliricidia sepium (Leguminosae) and Piper auritum (Piperaceae).

Susceptibility of human melanoma cells to oxidative stress including UVA radiation

Ultraviolet radiation, and in particular UVA (320–400 nm), induces significant oxidative stress to human skin. Ferritin and glutathione have been shown to be among the more important molecules within human skin cells providing protection against this damage, the presence of lower levels of these anti‐oxidants giving rise to increased cellular sensitivity to stress. We compared endogenous levels of ferritin and glutathione in human melanoma cells with normal human skin fibroblasts and keratinocytes, also the response of melanoma cells to oxidative stress with fibroblasts and keratinocytes. Ferritin levels were heterogenous in the untreated melanoma cell lines tested and remained the same following oxidative stress (UVA radiation) or hemin treatment. Epidermal keratinocytes were unaffected, as were the melanoma cell lines, but skin fibroblasts showed dose‐dependent ferritin depletion. Similar results were seen for glutathione alterations resulting from UVA radiation: melanoma cell lines and epidermal skin keratinocytes remained unchanged following UVA radiation, while skin fibroblasts showed dose‐dependent depletion. Our results show that human melanoma cells have low ferritin and glutathione levels, yet are resistant to oxidative stress.

Effects of Ultraviolet Radiation on Oxidative Stress Proteins in Human Skin in vivo

It is well known today, that exposure of human skin to solar light can result in short-tern responses, such as immune suppression, erythema and pigmentation, and long-term responses, such as carcinogenesis and premature aging (C.C. Cross et al., 1987). These deleterious biological effects are mainly due to the UVB portion of the sun spectrum, but recently it was shown, that wavelengths in the UVA range can produce similar biological effects by other mechanisms (L. A. Applegate et al., 1995; 3. Fuchs et al., 1989). These mechanisms involve endogenous chromophores such as quinons, steroids, flavins, free porphyrins and heme containing enzymes, which act as photosensitizers when induced by UVA irradiation. Reactive oxygen species (ROS), including singlet oxygen, superoxide radical anion, hydroxyl radical and hydrogen peroxide are produced during these photosensitized reactions in such a manner, that the proantioxidant balance can be overwhelmed (Y. Shindo et al., 1994). The resulting photo-oxidative stiess due to these harmful ROS is considered to play a major role in causing modifications of DNA, lipids, proteins and carbohydrates in human skin (K. Punnonen et al., 1991), which could lead to several cutaneous manefestations mentioned above.