Meta-analytic magic, ivermectin, and socially responsible reporting.

Some clinicians prescribe ivermectin for COVID-19 despite a lack of support from any credible South African professional body. They argue that when faced by clinical urgency, weak signals of efficacy should trigger action if harm is unlikely. Several recent reviews found an apparent mortality benefit by including studies at high risk of bias and with active rather than placebo controls. If these studies are discounted, the pooled mortality effect is no longer statistically significant, and evidence of benefit is very weak. Relying on this evidence could cause clinical harm if used to justify vaccine hesitancy. Clinicians remain responsible for ensuring that guidance they follow is both legitimate and reliable. In the ivermectin debate, evidence-based medicine (EBM) principles have largely been ignored under the guise thatin a pandemic the 'rules are different', probably to the detriment of vulnerable patients and certainly to the detriment of the profession's image. Medical schools and professional interest groups are responsible for transforming EBM from a taught but seldom-used tool into a process of lifelong learning, promoting a consistent call for evidence-based and unconflicted debate integral to clinical practice.

Tumor necrosis factor/cachectin-induced intravascular fibrin formation in meth A fibrosarcomas.

Recent studies have indicated that TNF can promote activation of the coagulation mechanism by modulating coagulant properties of endothelial cells. In this report, we demonstrate that infusion of low concentrations of TNF (3 micrograms/animal) into mice bearing meth A fibrosarcomas leads to localized fibrin deposition with formation of occlusive intravascular thrombi in close association with the endothelial cell surface. Studies with 125I-fibrinogen showed tenfold enhanced accumulation of radioactivity in tumor within 2 h after TNF infusion. Western blots of tumor extracts subjected to SDS-PAGE and visualized with a fibrin-specific mAb indicated that fibrin forms in the tumor after the TNF infusion. Electron microscopic studies demonstrated fibrin strands, based on the characteristic 21-nm periodicity, which appeared to be adherent to the endothelial cell surface. Further ultrastructural studies indicated that fibrin formation, first evident within 30 min of the TNF infusion, led to occlusive thrombi limited to the tumor vascular bed (i.e., not in the normal mouse vasculature) within 2 h and was associated with an 80% reduction in tumor perfusion based on studies with Evans blue. In view of previous work concerning TNF induction of endothelial cell procoagulant activity, the hypothesis that tumor cell products prime the response of endothelium to this cytokine was tested. Supernatants of cultured meth A fibrosarcomas obtained serum-free conditions, which had no intrinsic procoagulant activity, considerably enhanced tissue factor induction in endothelium in response to submaximal concentrations of TNF. The factor(s) in the tumor-conditioned medium appeared to be distinct from IL-1, fibroblast growth factor, IFN-gamma, TNF, endotoxin, TGF-alpha, and TGF-beta. These studies delineate a novel model of localized clot formation in which thrombosis is initiated by a pathophysiologic mediator, TNF, and provides an opportunity to examine mechanisms in the microenvironment directing clot formation to the tumor vascular bed.

Limited expression of heparan sulphate proteoglycans associated with Aß deposits in the APPswe/PS1dE9 mouse model for Alzheimer's disease.

AIMS: Alzheimer's disease (AD) is characterized by deposition of the amyloid beta (Aß) peptide in brain parenchyma and vasculature. Several proteins co-deposit with Aß, including heparan sulphate proteoglycans (HSPG). HSPG have been suggested to contribute to Aß aggregation and deposition, and may influence plaque formation and persistence by stimulating Aß fibrillization and by protecting Aß against degradation. Mouse models for AD, expressing the human amyloid precursor protein (APP), produce Aß deposits similar to humans. These models may be used to study disease pathology and to develop new therapeutic interventions. We aimed to investigate whether co-deposition of HSPG in AD brains can be replicated in the APPswe/PS1dE9 mouse model for AD and if a temporal association of HSPG with Aß exists. METHODS: We studied the co-deposition of several HSPG and of the glycosaminoglycan side chains of HSPG in the APPswe/PS1dE9 model at different ages by immunohistochemistry. RESULTS: We found that, although APPswe/PS1dE9 mice did develop severe Aß pathology with age, co-deposition of HS glycosaminoglycan chains and the various HSPG (agrin, perlecan and glypican-1) was scarce (<10-30% of the Aß deposits were stained). CONCLUSIONS: Our data suggest that the molecular composition of Aß deposits in the APPswe/PS1dE9 mouse, with respect to the several HSPG investigated in this study, does not accurately reflect the human situation. The near absence of HSPG in Aß deposits in this transgenic mouse model may, in turn, hinder the translation of preclinical intervention studies from mice to men.

Essential medicine selection during the COVID-19 pandemic: Enabling access in uncharted territory.

The COVID-19 pandemic requires urgent decisions regarding treatment policy in the face of rapidly evolving evidence. In response, the South African Essential Medicines List Committee established a subcommittee to systematically review and appraise emerging evidence, within very short timelines, in order to inform the National Department of Health COVID-19 treatment guidelines. To date, the subcommittee has reviewed 14 potential treatments, and made recommendations based on local context, feasibility, resource requirements and equity. Here we describe the rapid review and evidence-to-decision process, using remdesivir and dexamethasone as examples. Our experience is that conducting rapid reviews is a practical and efficient way to address medicine policy questions under pandemic conditions.

Micronodular transformation as a novel mechanism of VEGF-A-induced metastasis.

How and why tumors metastasize is still a matter of debate. The assumption is that mutations render tumor cells with a metastatic phenotype, enabling entrance in and transport through lymph or blood vessels. Distant outgrowth is thought to occur only in a suitable microenvironment (the seed and soil hypothesis). However, the anatomical location of most metastases in cancer patients suggests entrapment of tumor cells in the first microcapillary bed that is encountered. We here investigated how vascular endothelial growth factor-A (VEGF-A) attributes to the metastatic process. We describe here that VEGF-A enhances spontaneous metastasis by inducing intravasation of heterogeneous tumor cell clusters, surrounded by vessel wall elements, via an invasion-independent mechanism. These tumor clusters generate metastatic tissue embolisms in pulmonary arteries. Treatment of tumor-bearing mice with the antiangiogenic compound ZD6474 prevented the development of this metastatic phenotype. This work shows that tumors with high constitutive VEGF-A expression metastasize via the formation of tumor emboli and provides an alternative rationale for anti-VEGF-A therapy, namely to inhibit metastasis formation.

Data for indirect load case estimation of ice-induced moments from shaft line torque measurements.

During ice navigation, blade measurements of ice-induced moments on ship propellers, are challenged by the harsh operating environment. To overcome this problem, shaft line measurements are performed inboard, and the required propeller loads are subsequently estimated using a dynamic model and the solution of an inverse problem. The inverse problem is mathematically ill-posed and requires the determination of the ice-induced moment on the propeller blades from shaft line measurements. Full-scale torsional response data is presented as calculated from indirect strain measurements on the shaft line of a polar supply and research vessel. The vessel operated on a 68-day voyage between Cape Town and Antarctica and spent almost 11 days in sea ice with observed concentrations above 90% and a maximum thickness of 3 m. Data for five ice-induced load cases are presented, including the shaft torque from indirect measurements and the estimated ice-induced moment, which is obtained by solving an ill-posed inverse problem. The ice-induced moments on the propeller are obtained by approximating the drive-train as a viscously damped, elastic lumped mass model. The ice-induced moment is then determined through existing approaches to solving the ill-conditioned inverse problem. The lumped mass model is presented along with algorithms to solve the inverse problem, including truncated singular value decomposition, truncated generalized singular value decomposition and Tikhonov׳s method. The resulting time series data for the inversely calculated ice-induced moments is published to provide industry with load cases for ice-going propulsion design.

Complete response of melanoma-in-transit metastasis after isolated limb perfusion with tumor necrosis factor alpha and melphalan without massive tumor necrosis: a clinical and histopathological study of the delayed-type reaction pattern.

Treatment of stage IIIA/B melanoma patients by isolated limb perfusion (ILP) with a combination of tumor necrosis factor-alpha (TNF-alpha) and melphalan induces a complete response in 80-90% of the cases. The mechanism of tumor regression induced by the combination of TNF-alpha and melphalan is not precisely understood. Previous studies focused on the immediate (ie., within a few days) clinico-pathological changes after perfusion involving hemorrhagic necrosis. However, clinical data clearly indicate that complete tumor remission frequently requires a period of a few weeks to as much as months after ILP. Because the mechanism underlying this delayed-type reaction is completely unknown, we studied the clinico-pathological events in patients with such slowly regressing melanoma lesions. For this purpose, 94 biopsies of in-transit melanoma metastasis that were taken sequentially from 11 patients between 1 week and 9 months after ILP were analyzed by light and electron microscopy and immunohistochemistry. Clinical data included patient sex, age, anatomical localization and size of the tumor, and follow-up. All of the 11 patients ultimately responded to perfusion treatment (9 complete, 1 partial, 1 stable disease). Serial biopsies showed scattered individual tumor cell necrosis without hemorrhage. Most of the lesions with this delayed-type reaction pattern were less than 0.5 cm in diameter. They contained varying amounts of histologically viable-looking tumor cells and tumor-infiltrating melanophages. In addition, a marked but transient infiltrate of peritumoral eosinophils and moderate interstitial edema and dermal fibrosis were encountered. Only small numbers of lymphocytes were present. In comparison with the reaction pattern after treatment with melphalan alone, the delayed-type reaction pattern was similar but more intense. The scattered tumor cell necrosis in the latter type may be explained by a TNF-alpha-induced increase in permeability of the tumor vascular bed, which results in higher intratumoral concentrations of melphalan or in a prolongation of its effect. Subsequently, degenerated tumor cells are cleared by macrophages, and, finally, repair by fibrosis occurs. Because the immediate reaction type is evoked by hyperpermeability of the tumor vessels as well, quantitative differences seem to determine which reaction type ensues. We suggest that the extent of tumor vasculature that is sensitive to TNF-alpha determines the onset and histopathological pattern of tumor regression after ILP.

Hydroxylation of dehydroepiandrosterone in the eye lens.

Hydroxylation of the steroid hormone dehydroepiandrosterone in the calf lens is inhibited by carbon monoxide and stimulated by NADPH. The enzyme concerned was found to be membrane-bound. Although the enzyme resembles the liver mono-oxygenase system in these characteristics, the presence of cytochrome P-450 in the lens could not be proved by measuring a difference spectrum with carbon monoxide, probably because the concentration of the enzyme is too low. Preparations of purified lens fiber plasma membranes also hydroxylate dehydroepiandrosterone. This indicates that the fiber plasma membranes act as supports for enzyme complexes. In this respect they resemble cytoplasmic membranes and plasma membranes derived from other tissues. Cultured lens cell contain the hydroxylating enzyme, although its activity is dependent on the culture conditions used. It is striking that in lens fibers the enzyme which seems to convert dehydroepiandrosterone specifically occurs on the plasma membranes, whereas, for instance, in liver, hemoproteins localized on the endoplasmic reticulum, exert hydroxylation activity towards a variety of steroids. This suggests some regulatory role for dehydroepiandrosterone in lens growth and metabolism.

Serum IgA and gold-induced toxic effects in patients with rheumatoid arthritis.

Serum Immunoglobulin concentrations were measured prospectively in 25 patients with rheumatoid arthritis at months 0, 1, 3, 6, and 12 of aurothloglucose treatment. Substantial lowering of IgA and IgM levels was found at month 3 and thereafter, and of IgG at month 12 only. When patients in whom drug-induced toxic effects developed at any time during treatment (toxic group) were compared with those who did not (nontoxic group), serum levels of IgA and to a lesser degree of IgG, but not of IgM, were found to be substantially lower in the toxic than in the nontoxic group, both at the onset and during treatment, except for IgG at month 12. When measured at the moment of toxic effect, only igA, but not IgG and IgM, was substantially lower than in serum samples of patients without toxic effects at that moment. The serum IgA concentration in patients with rheumatoid arthritis seems to be related to whether or not aurothioglucose-induced toxic effects occur.

A new 180-kDa dermal endothelial cell activation antigen: in vitro and in situ characteristics.

PN-E2 is a monoclonal antibody generated against recombinant tumor necrosis factor-alpha (TNF-alpha)-treated human umbilical vein endothelial cells (EC). PN-E2 recognized a molecule with expression levels in vitro that could be downregulated by TNF, and in situ PN-E2 showed only weak reactivity with vascular EC in normal skin, as assessed by immunohistochemical staining. The expression of PN-E2 was considerably increased on EC in various pathologic skin lesions, including psoriasis, granulation tissue, and inflamed skin. PN-E2 antigen expression was analyzed in more detail in vitro on cultured EC and fibroblasts by use of enzyme-linked immunosorbent assay and fluorescence-activated cell sorter techniques. The expression level on human umbilical vein endothelial cells and capillary EC was, in contrast to the in situ immunohistologic findings, invariably high. On fibroblasts, a low expression was found. Incubation of the EC with recombinant TNF-alpha decreased expression by a factor of 2. Incubation of EC with recombinant interferon-gamma resulted in a twofold increase in PN-E2 antigen expression, whereas other cytokines [recombinant interleukin (rIL)-1 alpha, rIL-1 beta, rIL-4, rIL-6], lipopolysaccharide, or recombinant basic fibroblast growth factor had no effect. Immunoelectron microscopy of tissue specimens and EC preparations localized the antigen on the luminal membrane of the endothelium. Immunoprecipitation followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed a major band at 90 kDa and a minor band at 80 kDa under reducing conditions and bands of 180 and 400 kDa under non-reducing conditions. Molecular weight and expression patterns in vitro on EC after incubation with cytokines excluded most of the known endothelium-specific molecules, with the possible exception of endoglin (the 44G4 antigen). We conclude from our findings that this new antigen could be useful as a marker for endothelial activation in skin biopsy material.

Monoclonal antibody-defined human endothelial antigens as vascular markers.

A review is given of human endothelial antigens recognized by monoclonal antibodies and used as vascular markers. These antigens can be classified tentatively into two categories that partly overlap: 1) differentiation markers and 2) antigens involved in specific cellular functions. Monoclonal antibodies recognizing endothelial differentiation markers reacting with all types of human endothelium can be regarded as constitutive endothelial markers. Other differentiation markers have a restricted distribution that is associated with a subtype of endothelium. Although sensitivity of the markers is high in general, specificity for endothelium is not absolute, based on distribution studies in tissues or in cell lines. With the exception of PAL-E and EN-3/EN-4, it is not clear from the literature whether the antibodies also react with lymphatic endothelium. Immunohistochemical examination of other species indicate that only BW 200 is restricted to humans. Immunoelectron microscopy of microvascular cells in tissue specimens has revealed that the monoclonal antibodies recognizing differentiation antigens show different subcellular distribution patterns. PAL-E and BW 200 react with the luminal endothelial surface, in a local and diffuse pattern, respectively. Anti-Von Willebrand factor (i.e., Factor VIII-related ag) antibodies react with Weibel-Palade bodies but also with subendothelial structures. Applications of immunohistochemistry using monoclonal antibodies in diagnostic pathology include assessment of vascular invasion by cancer cells, and identification of endothelial neoplasms and related disorders. Because anti-Factor VIII-related antigen and BW 200 are applicable on formaldehyde-fixed and paraplast-embedded tissue, they are most suitable for histodiagnostic application. Immunohistochemistry using monoclonal antibodies recognizing endothelial antigens involved in specific cellular functions also may contribute to pathobiologic research on the characterization of blood-tissue barriers, e.g., in the tumor vascular bed.

Epidermal Langerhans cells contain intermediate-sized filaments of the vimentin type: an immunocytologic study.

In epidermal cell suspensions, prepared from healthy human and murine skin, Langerhans cells (LC) were identified by indirect immunofluorescence microscopy using monoclonal antibodies directed against human T6 and Ia markers, and against a murine Iak determinant, respectively. The cells were double-labeled on cytospin slides with antibodies against a cytoskeletal protein, either vimentin or keratin. In this way it was shown that epidermal LC contain intermediate-sized filaments of the vimentin type.

Phenotype of normal cutaneous microvasculature. Immunoelectron microscopic observations with emphasis on the differences between blood vessels and lymphatics.

The lymphatic system has been poorly characterized in comparison to the blood vessels. We investigated the expression of microvasculature markers in cutaneous lymphatics and blood microvessels in normal skin. Scrotal skin was chosen because of its high density of both types of microvessels. A pre-embedding peroxidase-conjugated immunoelectron microscopy technique was used, allowing both the visualization of the lymph and blood vessels and their immunohistochemical staining. The markers studied included endothelial antigens (recognized by PAL-E, EN-4, and von Willebrand factor/factor VIII-related antigen), structural molecules of the vascular wall (alpha-smooth muscle actin, heparan sulfate proteoglycan, collagen type IV), and adhesion molecules (endothelial leukocyte adhesion molecule-1 [E-selectin], intercellular adhesion molecule-1 [ICAM-1], platelet endothelial adhesion molecule-1 [PECAM-1], vascular cell adhesion molecule-1 [VCAM-1]). It is shown that lymphatics of normal skin are phenotypically different from blood microvasculature, only weakly expressing endothelial markers (EN-4+, von Willebrand factor/factor VIII-related antigen +/-, PAL-E-), mural markers (alpha-smooth muscle actin-, heparan sulfate proteoglycan-, collagen type IV+) and do not express the studied adhesion molecules except PE-CAM-1 (E-selectin-, ICAM-1-, PECAM-1+, VCAM-1-). The results were substantiated by a double-labeling immunoelectron microscopic technique, which facilitates detection and assessment of microvascular segments. By this technique, collagen type IV, recognized by a peroxidase-labeled 2nd antibody, stains the basal lamina by a linear pattern, whereas a second optional epitope is visualized as grains by a silver-enhanced ultra-small gold-conjugated antibody. Our study shows that not only morphology but also antigenic phenotype of lymphatics differs significantly from blood vessels.

4-Aminopyridine is superior to 3,4-diaminopyridine in the treatment of patients with multiple sclerosis.

OBJECTIVE: To compare the efficacy and toxicity of 4-aminopyridine and 3,4-diaminopyridine in patients with multiple sclerosis. DESIGN: Intervention study with a before-after design and a randomized, double-blind, crossover design. SETTING: University referral center. PATIENTS: Twenty-four patients with definite multiple sclerosis who had been treated in a previous clinical trial with 4-aminopyridine. INTERVENTIONS: Nonresponders to treatment with 4-aminopyridine (14 patients) were treated with 3,4-diaminopyridine in a 4-week, open-label trial with doses up to 1.0 mg/kg of body weight (before-after design). Responders to treatment with 4-aminopyridine (10 patients) participated in a comparative study of 6 weeks' duration with 4-aminopyridine and 3,4-diaminopyridine according to a randomized, double-blind, double-crossover design. MAIN OUTCOME MEASURES: Neurophysiologic variables for nonresponders, neurologic functions and symptoms on a visual analogue scale for responders, and side effects for both groups. RESULTS: Toxicity profiles of 4-aminopyridine and 3,4-diaminopyridine were different, and systemic tolerability was reduced for 3,4-diaminopyridine. 4-Aminopyridine was more effective than 3,4-diaminopyridine, especially for ambulation, fatigue, and overall daily functioning. CONCLUSION: Our data suggest that, concerning both efficacy and side effects, 4-aminopyridine is superior to 3,4-diaminopyridine in the treatment of patients with multiple sclerosis.

Procedure for the isolation of mouse anti H-2 antibodies from complex alloantisera.

B6AF1-anti-B10.D2 ascites fluid was incubated with donor strain red blood cells to absorb the anti-H-2 antibodies. The antibody-coated RBC were then lysed, and from the ghosts the antibodies were eluted. This anti-H-2 preparation was further purified by affinity chromatography, using an immobilized anti IgG2 antiserum. Thus a pure anti-H-2 IgG2 was obtained for in vitro and in vivo testing.

An improved sensitive and simple microassay of mouse complement.

A simple and fast hemolytic microassay was developed for the determination of classical pathway complement activity in mouse serum. The assay is based on hemolysis of sheep red blood cells (SRBC) that are sensitized with polyclonal mouse antibodies. The degree of hemolysis was measured in the reaction supernatants by photometric reading in an ELISA plate scanner at 405 nm wavelength. It was found that some batches of unpurified mouse anti-SRBC antibodies gave insufficient hemolysis. Analysis of two antibody preparations indicated that this might be caused by anti-complementary factors in the ascites fluid, and by an excess of non-complement fixing IgG1 antibodies. For optimal and standardized results, removal of anticomplementary factors and enrichment for complement fixing IgG2 antibodies was required and was achieved using protein A purified anti-SRBC IgG. In our assay it is possible to determine CH50 titers in triplicate in 80 microliters samples of individual mouse sera with high sensitivity. Using this rapid one-step method large numbers of tests could be performed in 1 day.

In vivo activity of an H-2 alloantiserum purified by affinity chromatography on transplantation antigens.

Transplantation antigens were isolated from murine tissue by solubilization with NP-40 and purification on a lentil lectin-Sepharose column. The glycoproteins eluted from the lectin column were coupled to CNBr-activated Sepharose to prepare a specific immunoadsorbent. Intact histocompatibility antigens were demonstrated on the carrier particles in a cytotoxic inhibition assay with specific antiserum and spleen lymphocytes. Adsorption and subsequent elution of alloantiserum directed against the coupled antigens yielded a 30-fold purified product. The presence of both active K and I region products on the column was demonstrated by the in vivo activities of the alloantibodies eluted from the coupled molecules. Both acute rejection, a function of H-2K antibodies, and passive enhancement, a function of Ia antibodies, could be induced by administration of the eluate to recipients of skin allografts. These in vivo results show that the adsorbent can be used for preparative purposes.

Expression of donor class I major histocompatibility antigens on the vascular endothelium of mouse skin allografts.

The expression of a class I MHC antigen on the vascular endothelium of mouse skin allografts was assessed by in vivo uptake of radiolabeled monoclonal anti-class-I antibody in the grafts after i.v. injection into the recipients. Endothelial localization of the bound antibodies was demonstrated via double-labeling immunofluorescence microscopy using factor-VIII-related antigen as a marker for endothelial cells. Treatment of recipients with cyclosporine was accompanied by low levels of class I antigen expression in the grafts, and similarly low levels were measured in grafts carried by nude recipients in the complete absence of rejection. Withdrawal of immunosuppressive therapy was followed by an increased class I antigen expression in the donor skin. An increase was also observed in skin grafts undergoing first-set rejection. We conclude that the expression of class I antigens on the capillary endothelium of mouse skin allografts in vivo is variable and is under influence of the immune status of the recipient.

Lack of lymphangiogenesis despite coexpression of VEGF-C and its receptor Flt-4 in uveal melanoma.

PURPOSE: Because lymphatic vessels are absent from the normal eye and because uveal melanomas are presumed to spread by a hematogenous route in the absence of tumor exposure to conjunctival lymphatics, this study was undertaken to investigate the presence of lymphatic vessels in primary uveal melanomas. METHODS: The presence of lymphatics in 2 control eyes and in 33 primary uveal, 10 primary cutaneous, and 3 metastatic cutaneous melanomas was evaluated by using a double-immunostaining protocol that differentially highlights blood and lymphatic vasculature. In addition, 14 uveal melanomas were immunostained for the lymphatic growth factor vascular endothelial growth factor (VEGF)-C (with anti-VEGF-C polyclonal antibodies [pAbs]), its receptors Flt-4 (with monoclonal antibody [mAb] 9D9) and KDR (with anti-KDR mAb [Clone KDR-2]), and the hemangiogenic factor VEGF-A (with anti-VEGF pAbs). RESULTS: Lymphatics were not detected in normal eyes or in uveal melanoma. As a consequence, signs of lymphangiogenesis were not present. There was coexpression of VEGF-C with Flt-4 and KDR in 6 (43%) of the 14 melanomas. Staining for VEGF-A was completely negative in 25 uveal melanomas analyzed. CONCLUSIONS: The strictly hematogenous metastasis of primary uveal melanomas is explained by the absence of lymphatics in and around the tumor. The current data suggest that, in the presence of endothelial Flt-4, VEGF-C expression is not sufficient to induce lymphangiogenesis from preexisting blood vessels in human cancer.

Measurement of tissue factor messenger RNA levels in human endothelial cells by a quantitative RT-PCR assay.

We have developed a sensitive and quantitative RT-PCR assay for the determination of tissue factor (TF) mRNA levels in human cells. An in vitro synthesized internal standard RNA was used to correct for differences in reverse transcription or amplification of various RNA samples. The PCR products were quantitated by hybridization. The sensitivity was such that less than 0.2 microgram of total endothelial RNA sufficed to measure its TF mRNA content. The RT-PCR assay was used to determine TF mRNA levels in endothelial cells treated with a factor from human melanoma cells and/or TNF. In this way the amount of TF mRNA could be induced to a level that was at least 80-fold higher than that in non-induced cells. This increase was in the same order of magnitude as the induction of measured TF activity.