Comparison of neoadjuvant chemotherapy versus upfront surgery with or without chemotherapy for patients with clinical stage III esophageal squamous cell carcinoma.

Neoadjuvant chemotherapy (NAC) and chemoradiotherapy have been shown to extend postoperative survival, and preoperative therapy followed by esophagectomy has become the standard treatment worldwide for patients with esophageal squamous cell carcinoma (ESCC). The Japan Clinical Oncology Group 9907 study showed that NAC significantly extended survival in advanced ESCC, but the survival benefit for patients with clinical stage III disease remains to be elucidated. We compared the survival rates of NAC and upfront surgery in patients with clinical stage III ESCC. Consecutive patients histologically diagnosed as clinical stage III (excluding cT4) ESCC were eligible for this retrospective study. Between September 2002 and April 2007, upfront transthoracic esophagectomy was performed initially and, for patients with positive lymph node (LN) metastasis in a resected specimen, adjuvant chemotherapy using cisplatin and 5-fluororouracil every 3 weeks for two cycles was administered (Upfront surgery group). Since May 2007, a NAC regimen used as adjuvant chemotherapy followed by transthoracic esophagectomy has been administered as the standard treatment in our institution (NAC group). Patient characteristics, clinicopathological factors, treatment outcomes, post-treatment recurrence, and overall survival (OS) were compared between the NAC and upfront surgery groups. Fifty-one and 55 patients were included in the NAC and upfront surgery groups, respectively. The R0 resection rate was significantly lower in the NAC group than in the upfront surgery group (upfront surgery, 98%; NAC, 76%; P = 0.003). In the upfront surgery group, of 49 patients who underwent R0 resection and pathologically positive for LN metastasis, 22 (45%) received adjuvant chemotherapy. In the NAC group, 49 (96%) of 51 patients completed two cycles of NAC. In survival analysis, no significant difference in OS was observed between the NAC and upfront surgery groups (NAC: 5-year OS, 43.8%; upfront surgery: 5-year overall surgery, 57.5%; P = 0.167). Patients who underwent R0 resection showed significantly longer OS than did those who underwent R1, R2, or no resection (P = 0.001). In multivariate analysis using age, perioperative chemotherapy, depth of invasion, LN metastasis, surgical radicality, postoperative pneumonia, and anastomotic leakage as covariates, LN metastasis [cN2: hazard ratio (HR), 1.389; P = 0.309; cN3: HR, 16.019; P = 0.012] and surgical radicality (R1: HR, 3.949; P = 0.009; R2 or no resection: HR, 2.912; P = 0.022) were shown to be significant independent prognostic factors. In clinical stage III ESCC patients, no significant difference in OS was observed between NAC and upfront surgery. Although potential patient selection bias might be a factor in this retrospective analysis, the noncurative resection rate was higher after NAC than after upfront surgery. The survival benefit of more intensive NAC needs to be further evaluated.

Delayed-onset ataxia in mice lacking alpha -tocopherol transfer protein: model for neuronal degeneration caused by chronic oxidative stress.

alpha-Tocopherol transfer protein (alpha-TTP) maintains the concentration of serum alpha-tocopherol (vitamin E), one of the most potent fat-soluble antioxidants, by facilitating alpha-tocopherol export from the liver. Mutations of the alpha-TTP gene are linked to ataxia with isolated vitamin E deficiency (AVED). We produced a model mouse of AVED by deleting the alpha-TTP gene, which showed ataxia and retinal degeneration after 1 year of age. Because the brain alpha-TTP functions in maintaining alpha-tocopherol levels in the brain, alpha-tocopherol was completely depleted in the alpha-TTP(-/-) mouse brain, and the neurological phenotype of alpha-TTP(-/-) mice is much more severe than that of wild-type mice when maintained on an alpha-tocopherol-deficient diet. Lipid peroxidation in alpha-TTP(-/-) mice brains showed a significant increase, especially in degenerating neurons. alpha-Tocopherol supplementation suppressed lipid peroxidation and almost completely prevented the development of neurological symptoms. This therapy almost completely corrects the abnormalities in a mouse model of human neurodegenerative disease. Moreover, alpha-TTP(-/-) mice may prove to be excellent animal models of delayed onset, slowly progressive neuronal degeneration caused by chronic oxidative stress.

C8/119S mutation of major mite allergen Derf-2 leads to degenerate secondary structure and molecular polymerization and induces potent and exclusive Th1 cell differentiation.

Hyposensitization therapy for atopic diseases has been conducted for decades but suffered from many problems including anaphylactic reactions. We previously developed a mutant protein of the major mite allergen Derf-2, C8/119S, which showed reduced binding to IgE. The C8/119S mutant was shown to exhibit more efficient hyposensitizing effect than Derf-2 in the animal model of allergic bronchial asthma. In the present study, we indicate that C8/119S exhibits markedly augmented immunogenicity for the proliferation of Derf-2-specific human T cells and T cell clones irrespective of the epitope specificity as compared with Derf-2. Furthermore, C8/119S has induced potent and almost exclusive differentiation of Th1 cells from the peripheral blood of atopic patients in vitro. Neither Ag dosage effect nor absence of B cell-mediated Ag presentation could fully account for these effects. C8/119S has been indicated to lose the characteristic beta-barrel structure as judged by circular dichroism spectroscopic analysis and to polymerize solubly in physiological condition. Heating of Derf-2 also caused less stable molecular aggregation, but it hardly affected the secondary structure and failed to induce such a polarity toward the Th1 cell differentiation. These results have indicated that the degenerate secondary structure of C8/119S leading to stable molecular polymerization is primarily responsible for the marked increase in T cell-immunogenicity and the induction of exclusive Th1 cell differentiation in atopic patients. It has been suggested strongly that the recombinant C8/119S protein can provide an effective Ag with the least risk of anaphylaxis for allergen immunotherapy against house dust mite in human.

Postmortem study of ataxia with retinitis pigmentosa by mutation of the alpha-tocopherol transfer protein gene.

A new syndrome of ataxia and retinitis pigmentosa with vitamin E deficiency caused by the missense mutation of alpha-tocopherol transfer protein (alpha-TTP) gene was recently proposed. After studying the first postmortem case with this mutation pathologically and biochemically, whether the symptoms can be treated by supplementation of vitamin E or not is discussed. The major pathological findings were retinal atrophy; severe dying back-type degeneration of the posterior column; and massive accumulation of lipofuscin in neurons including dorsal root ganglion (DRG) cells, which were almost identical to those in vitamin E deficient animals and patients with fat malabsorption. Also, mild loss of Purkinje cells was noted. Because robust expression of alpha-TTP was detected in the cerebellum as well as in the liver and the tissue concentration of vitamin E in the cerebellum was still low even after oral supplementation, the mild Purkinje cell loss might be related to the mutant alpha-TTP in the cerebellum. By contrast, in the DRG, thought to be mainly responsible for ataxia, no expression of alpha-TTP was detected, and the tissue concentration of vitamin E increased to normal after supplementation. It is therefore considered that oral supplementation of vitamin E should effectively counteract the progression of ataxia.

The striking effect of hyperbaric oxygenation therapy in the management of chronic idiopathic intestinal pseudo-obstruction.

Chronic idiopathic intestinal pseudo-obstruction is one of the disorders that is most refractory to medical and surgical treatment. Even when patients are given nutritional support, including total parenteral nutrition, obstructive symptoms seldom disappear. We report a case of chronic idiopathic intestinal pseudo-obstruction, due to myopathy, in which hyperbaric oxygenation therapy was strikingly effective. The presence of myopathy was histologically confirmed on the surgically resected jejunal specimen. Hyperbaric oxygenation resulted not only in relief of the patient's obstructive symptoms but also in a rapid decrease of abnormally accumulated intestinal gas. At last, he could resume oral intake without any critical adverse effects. These observations strongly suggest that hyperbaric oxygenation can be an effective therapy in the management of chronic idiopathic intestinal pseudo-obstruction.

[Surgical treatment of N2 involved non-small cell lung cancer--the systematic extended lymph node dissection based on the regional lymphatic drainage].

From the study on regional lymphtic drainage, we have decided the extent of lymphadenectomy as follows; a) For the left lung cancer and the right upper lobe primary, systematic bilateral mediastinal dissection (R3 alpha) through a median sternotomy, b) For the cases with the highest mediastinal node involvement, lower half of radical neck dissection (R3 gamma) through a cervical collar incision in addition to a). The cN diagnosis by CT interpretation and pN diagnosis were compared. The under estimated rates of N were 32% of 137 patients with the left lung primary. 46 patients with pN2(+) included 14 patients (31%) with pN3 disease. As for the right upper lobe primary, 17 patients with pN2(+) included 13 patients (76%) with pN3 disease. Postoperative survival rates calculated with Kaplan-Meiermethod; 1) The five-year survival rates were 43% of 46 patients with pT1-3 N2-3 of the left lung primary. 2) As for the right upper lobe primary, the two-year survival rates were 51% of 17 patients with pT1-4 N2-3. 3) The three-year survival rates of 26 patients with pN3 gamma diagnosed as cN0-3 alpha preoperatively were 41%. These systematic extended dissection (R3 alpha, R3 gamma) would bring better prognosis in the patients with pN2-3 disease.

The inhibitory effect of glabridin from licorice extracts on melanogenesis and inflammation.

Glabridin is the main ingredient in hydrophobic fraction of licorice extract affecting on skins. In this study, we investigated inhibitory effects of glabridin on melanogenesis and inflammation using cultured B16 murine melanoma cells and guinea pig skins. The results indicated that glabridin inhibits tyrosinase activity of these cells at concentrations of 0.1 to 1.0 microg/ml and had no detectable effect on their DNA synthesis. Combined analysis of SDS-polyacrylamide gel electrophoresis and DOPA staining on the large granule fraction of these cells disclosed that glabridin decreased specifically the activities of T1 and T3 tyrosinase isozymes. It was also shown that UVB-induced pigmentation and erythema in the skins of guinea pigs were inhibited by topical applications of 0.5% glabridin. Anti-inflammatory effects of glabridin in vitro were also shown by its inhibition of superoxide anion productions and cyclooxygenase activities. These data indicated that glabridin is a unique compound possessing more than one function; not only the inhibition of melanogenesis but also the inhibition of inflammation in the skins. By replacing each of hydroxyl groups of glabridin with others, it was revealed that the inhibitory effect of 2'-O-ethyl glabridin was significantly stronger than that of 4'-O-ethyl-glabridin on melanin synthesis in cultured B16 cells at the concentration of 1.0 mg/ml. With replacement of both of two hydroxyl groups, the inhibitory effect was totally lost. Based on these data, we concluded that two hydroxyl groups of glabridin are important for the inhibition of melanin synthesis and that the hydroxyl group at the 4' position of this compound is more closely related to melanin synthesis.

Distribution of nociceptive neurons in the ventrobasal complex of macaque thalamus.

In urethane-chloralose anesthetized Japanese macaques, the distribution of nociceptive neurons within the thalamic ventrobasal (VB) complex was studied. Nociceptive specific (NS) and wide dynamic range (WDR) neurons were found in the periphery of the contralateral integument compartment of the VB complex. Thus, they formed a shell at the perimeter of this compartment with a somatotopic organization. The compartment consisted of large parts of nucleus ventralis posteromedialis (VPM) and nucleus ventralis posterolateralis, pars caudalis (VPLc). NS neurons were located more caudally than WDR neurons. In the NS zone of VPM, the forehead was represented caudally, and oral structures rostrally. In the ventral NS zone of VPM, there was a sequential representation of the tongue, gum and mandibular skin from the medial to the lateral edge. The hand was represented medially in the NS zone of VPLc, and its representation dominated in the rostral NS zone. There was a sequential representation of the cervical, thoracic, lumbar, sacral and caudal segments mediolaterally along the dorsal VPLc. In the medial half of ventral NS zone of VPLc, the upper body half was represented, and in the lateral half, the lower body half. The foot was represented at or near the medial edge of lateral half. In the rostral WDR zone, the trunk and peripheral face were represented.

Anastomotic recurrence after curative resection of a transverse colon carcinoma: a case report.

We report a case of anastomotic recurrence after curative surgery for transverse colon cancer in a 53-year-old man. The recurrence was first detected as a submucosal tumor 1.3 cm in diameter, located on the suture line, during an annual follow-up barium enema and colonoscopy. A repeat examination 3 months later showed the lesion to be a typical colon cancer, 2.5 cm in size, with a large ulcerated area. Right hemicolectomy was performed with curative intent. Anastomotic recurrence is much rarer after colonic resection than after anterior resection. This was the first time that we had detected a recurrent lesion as a submucosal tumor during annual follow-up examination.

Abdominal actinomycosis: barium enema and computed tomography findings.

A case of abdominal actinomycosis is described in a woman with recurrent right lower abdominal pain and low-grade fever without history of appendectomy. Past history included the use of an intrauterine device (IUD) until 10 years before manifestation of these symptoms. We followed up the patient, via diagnostic imaging, for 7 months. On initial barium enema, a polypoid lesion was visualized at the bottom of the cecum and there was constriction of the sigmoid colon; the appendix was not seen. Seven months later, poor extension at the cecum, severe constriction in the sigmoid colon, and narrowing of the terminal ileum were also visualized. On computed tomography (CT), the lesion was initially localized only in the ileocecal region adjacent to the sigmoid colon. After 7 months, the lesion had infiltrated adjacent anatomic components and showed direct infiltration of the pelvic space. Differential diagnosis was difficult, as it was not obvious whether this was a pelvic abscess due to inflammation or appendiceal carcinoma. Laparotomy was performed. Macroscopically, the lesion was not limited to the ileocecal region, but involved the right ureter, tubes the Fallopian and ovary, bladder, psoas muscle, and abdominal wall. Pathology findings showed, chronic inflammatory tissue with evidence of actinomycosis. Although previous reports have described a lack of specific findings in this disease. When actinomycosis is suspected, CT is recommended to define its extent.

Establishment of an in vitro assay system for screening hepatitis C virus protease inhibitors using high performance liquid chromatography.

The hepatitis C virus (HCV) genome contains the code for a conserved, serine-type protease, called NS3, for the processing of the non-structural protein region of the viral polyproteins. Furthermore, a related protein, NS4A, is an effector or cofactor of NS3 protease activity in the cleavage of NS3-4A, NS4A-4B, NS4B-5A and NS5A-5B junctions. To establish an in vitro assay system for the screening of those enzyme inhibitors that inhibit the protease NS3-4A, we prepared a maltose-binding protein-NS3-NS4A fusion protein and a synthetic peptide substrate that mimics the NS5A-5B junction. Cleavage of the synthetic peptide was analyzed by reversed-phase high performance liquid chromatography (HPLC). We showed that the enzymatic activity of the NS3-NS4A fusion protein was enhanced in comparison to the NS3 protein alone. The assay conditions for optimum NS3-4A protease activity were determined to be pH 7.6 and 37 degrees C. In addition, we evaluated several protease inhibitors using the same HPLC assay system. The activity of HCV protease NS3-4A was inhibited by 2714.4 microM diisopropyl fluorophosphate, 270.8 microM N-tosyl-L-lysyl chloromethyl ketone, and 825.5 microM chymostatin. The results of the present study indicated that the synthetic peptide substrate and HPLC assay system are suitable for studying HCV protease activity and may facilitate the development of anti-HCV therapeutic reagents.

Repression of acute gastric mucosal lesions by antioxidant-containing fraction from fermented products of okara (bean-curd residue).

A crude antioxidant preparation from fermented okara (NTX) was examined for its protection against the pathogenesis of gastric ulcer in water-immersed rats. The areas of gastric mucosal lesions as well as the levels of thiobarbituric acid-reactive substances, prostaglandin E2 and hyaluronic acid in the gastric mucosa were measured in relation to the time elapsed after the imposition of stress. Comparison with those parameters in alpha-tocopherol-treated groups revealed that NTX exerted an anti-inflammatory effect on gastric injury, probably by functioning as a free radical scavenger.

Colorimetric assay system for screening antiviral compounds against hepatitis B virus.

A highly sensitive, rapid, and accurate assay system was developed for the in vitro evaluation of anti-hepatitis B virus (anti-HBV) agents. Chronic HBV-producing HB611 cells were used in combination with immunoaffinity purification, polymerase chain reaction (PCR), and hybrid capture detection. HB611 cells were incubated with putative anti-HBV agents for 7 days in 96-well microtiter plates. HBV was purified from HB611 cell culture media using immunoaffinity purification. The HBV DNA was extracted, amplified with PCR, and assayed using a hybrid capture colorimetric method. This assay provided quantitative detection of extracellular HBV DNA from 25 microliters of cell culture media. Using the colorimetric method, we found that 50% effective concentration levels of several known anti-HBV agents (HPMPA, PMEDAP, PMEA and others) were similar to those reported in studies using Southern blot analysis. These results demonstrate that this new and easily automated colorimetric assay system can be used for the rapid and accurate assessment of anti-HBV compound selectivity.

[Effect of direct current therapy with chemotherapy in 7,12-dimethylbenz[alpha] anthracene (DMBA)-induced rat mammary cancer].

To improve the quality of life of breast cancer patients with local recurrence, we investigated the effect of direct current therapy in combination with chemotherapy. In this study we chose 3mA of direct current and 1mg/kg of CDDP as optimum based upon a prior experimental study. Rats with mammary cancer induced by 7,12-dimethylbenz [alpha] anthracene (DMBA) were divided into four groups based upon therapeutic methods: direct current therapy alone (Group DC), chemotherapy alone (Group CT), direct current therapy with chemotherapy (Group DC+CT) and no treatment (control). The percentage of tumor area 6 weeks after therapy was 30.8% of the metropeutic size in the Group DC+CT, 46.8% in the Group DC, 107% in the Group CT and 267% in the control (Group DC+CT vs the other groups: p < 0.05). The response rate in the Group DC+CT (100%) was significantly higher than that in the other three groups (chi 2-test, p < 0.05). The platinum concentration ratio of both tumor/serum (p < 0.01) and tumor/kidney (p < 0.05) in the Group DC+CT was significantly higher than that in the Group CT. We conclude that direct current therapy in combination with chemotherapy strongly reduces mammary tumor size of rats.

A screening system for antiviral compounds against herpes simplex virus type 1 using the MTT method with L929 cells.

We developed the improved MTT assay system for in vitro evaluation of anti-HSV-1 agents using L929 cells derived from mouse connective tissue. This assay system provides results in 4 days using a 96-well microplate. The EC50 values of several anti-HSV agents (ACV, BVaraU, and others) were found to be similar to those obtained by the plaque reduction method using MRC-5 cells. The present MTT assay is rapid, accurate, and may be useful as an automatic screening system for evaluation of anti-HSV-1 compounds.

Effects of dietary protein on the induction of DNA synthesis and expression of growth-related genes in liver and kidney of growing rats.

To investigate molecular mechanisms of growth control by protein nutrition, we examined whether nutritive quality of protein affects the induction of DNA synthesis in liver and kidney of growing rats in relation to expression of growth-related genes such as c-myc, c-fos, c-Ha-ras, and ornithine decarboxylase (ODC). Rats were adapted to 2-h meal feeding schedule at first with laboratory chow for 10 days and then with a protein-free diet for 3 days prior to experiments. When protein-free diet was fed to the rats, the levels of c-myc, ODC and c-Ha-ras mRNAs increased in the liver within 2 days. However, substantial changes in the levels of those mRNAs were not observed in the kidney. The level of c-fos mRNA in these tissues was too low to detect by our method. Feeding of casein diet to rats that had been maintained on protein-free diet for 3 days caused a rapid decrease in the level of c-myc mRNA and induced DNA synthesis in the liver. On the other hand, zein diet, which lacks tryptophan and lysine, did not lower the c-myc mRNA level nor induced DNA synthesis in the liver. However, if zein diet was supplemented with tryptophan and lysine, a decrease in c-myc mRNA level and an induction of DNA synthesis were observed. The levels of ODC and c-Ha-ras mRNAs were not changed by feeding of casein or zein diet. Neither casein nor zein induced DNA synthesis and changed the levels of the mRNA in the kidney. The amount of food intake during the 2-h feeding period was not different among the diets. These results suggest that the liver cells are arrested in G1 phase during the feeding of protein-free diet and good quality of protein is required to progress the cell cycle to enter S phase.

Differential inhibitory mechanisms in VPL versus intralaminar nociceptive neurons of the cat: I. Effects of periaqueductal gray stimulation.

Nociceptive thalamic units receiving afferent input from the greater splanchnic nerve (SPL) were recorded from the nucleus ventralis posterolatealis (VPL) and intralaminar nuclei in urethane-chloralose anesthetized cats. The effects of stimulating the periaqueductal gray (PAG), or the nucleus raphe dorsalis (NRD) on responses of nociceptive thalamic units were investigated. Forty-eight nociceptive specific (NS) and 20 wide dynamic range (WDR) units with SPL input were found in the shell region of the caudal VPL. Following electrical stimulation of either the ventral PAG or the NRD, responses to SPL input were inhibited in all NS and WDR units tested. Responses of these units to electrical stimulation of spinothalamic tract fibers in the ventrolateral funiculus (VLF) were also inhibited following the PAG/NRD stimulation. These results suggest that PAG/NRD stimulation-produced inhibition of both NS and WDR units may be partially mediated by an ascending antinociceptive mechanism. Intralaminar nociceptive units with SPL input were found in the nuclei centralis lateralis (CL), paracentralis (Pc), and parafascicularis (Pf). The effects of conditioning electrical stimulation of either the ventral PAG or the NRD on responses of intralaminar nociceptive units were studied. Of 113 intralaminar nociceptive units studied, 68 units were unaffected, 23 units were excited and 22 units were inhibited following the conditioning stimulation. In the units in which responses to SPL stimulation were inhibited, responses elicited by electrical stimulation of the mesencephalic reticular formation (MRF) were also inhibited. These data suggest that although there is an ascending inhibitory pathway from PAG/NRD to intralaminar nuclei, this system is far less potent compared with the ascending inhibitory system acting upon the VPL.

Differential inhibitory mechanisms in VPL versus intralaminar nociceptive neurons of the cat: II. Effects of systemic morphine and CCK.

In urethane-chloralose anesthetized cats, the effects of intravenous morphine on responses of thalamic nociceptive units were studied. In both nociceptive specific (NS) and wide dynamic range (WDR) units recorded from the nucleus ventralis posterolateralis (VPL), intravenous morphine suppressed unit responses to the greater splanchnic nerve (SPL) stimulation, but had little effect on responses to stimulation of spinothalamic tract fibers in the ventrolateral funiculus. In nociceptive units recorded from nuclei centralis lateralis (CL) and parafascicularis (Pf) of the intralaminar nuclei, intravenous morphine suppressed responses to stimulation of the mesencephalic reticular formation as well as to SPL stimulation. Intravenous cholecystokinin (CCK) antagonized the suppressive action of morphine on responses of VPL units, but did not antagonize the suppressive action of morphine on responses of intralaminar units. The results suggest that intravenous morphine inhibits synaptic transmission of nociceptive impulses in the intralaminar nuclei as well as in the spinal cord, but not in the VPL, and that CCK antagonizes the antinociceptive action of morphine in the spinal cord, but not in the intralaminar nuclei.

[Fundamental and clinical studies of SY5555 in pediatrics].

We assessed the in vitro antimicrobial activity and the clinical efficacy and safety of SY5555 in the field of pediatrics. The results obtained are summarized below. 1. In vitro antibacterial activities of SY5555 against 52 clinical isolates were compared with those of clavulanic acid/amoxicillin (CVA/AMPC), cefotiam (CTM), cefpodoxime (CPDX), cefaclor (CCL) and cefdinir (CFDN). Against Gram-positive bacteria, including Staphylococcus aureus, Streptococcus pneumoniae and Streptococcus pyogenes, SY5555 displayed antimicrobial activities superior or nearly equivalent to those of the reference agents used in the study. In cases of Gram-negative bacteria, the antimicrobial activity of SY5555 against Haemophilus influenzae was inferior to those of CPDX and CFDN. Against Klebsiella pneumoniae, the antimicrobial activity of SY5555 was less potent than that of CPDX. 2. Forty-seven children with infectious diseases were treated with SY5555 dry syrup (powder dissolved just before use). The clinical results were excellent in 24 and good in 16, with an efficacy rate of 85.1%. 3. Bacteriological screening identified 30 pathogenic organisms, and the eradication rate was 76.7%. 4. Side effects consisted of diarrhea in 12.5% (6 cases), loose stools in 4.2% (2 cases) and urticaria in 2.1% (1 case) of the patients. The only abnormal laboratory test value observed was an increase in eosinophil count in one child. 5. The palatability of SY5555 dry syrup was very good; it was very easily ingestable or easily ingestable by 32 of the 48 children. From the above results, SY5555 dry syrup appears to be a useful drug with a preferable safety profile in the treatment of pediatric patients with infectious diseases.

Molecular cloning of a novel human cDNA encoding a zinc finger protein that binds to the interleukin-3 promoter.

The CT/GC-rich region (-76 to -47) is one transcriptional regulatory region of the interleukin-3 (IL-3) gene which confers basic transcriptional activity and responds to trans-activation by human T-cell leukemia virus type I-encoded Tax. We isolated three types of cDNAs encoding Cys2/His2-type zinc finger proteins that bind to this region. Two were identical to known transcription factors, EGR1 and EGR2, and the other clone, named DB1, encoded a novel protein of 516 amino acids with six zinc finger motifs. DB1 mRNA was present in human tissues, ubiquitously. Two constitutive transcripts of 4.0 and 4.8 kb in length were present in Jurkat cells. Electrophoretic mobility shift assay, with specific antibodies, showed that DB1 constitutively binds to this region whereas EGR1 binds in a T-cell activation-dependent manner. Overexpression of DB1 in Jurkat cells had no detectable effect on the transcription activity of the IL-3 promoter, in a transient-transfection assay. EGR1 and EGR2 increased IL-3 promoter activity when the transfected cells were stimulated with phorbol-12-myristate-13-acetate and A23187. When DB1 was cotransfected with a Tax expression vector, transcription activity of the IL-3 promoter induced by Tax was significantly increased, while EGR1 and EGR2 were without effect. These results suggest that EGR1 has a role in inducible transcription of the IL-3 gene, while DB1 sustains basal transcriptional activity and also cooperates with Tax to activate the IL-3 promoter.