Identification of recognition sites for myc/max/mxd network proteins by a whole human chromosome 19 selection strategy.

In this study, we have identified 20 human sequences containing Myc network binding sites in a library from the whole human chromosome 19. We demonstrated binding of the Max protein to these sequences both in vitro and in vivo. The majority of the identified sequences contained one or several CACGTG or CATGTG E-boxes. Several of these sites were located within introns or in their vicinity and the corresponding genes were found to be up- or down-regulated in differentiating HL-60 cells. Our data show the proof of principle for using this strategy in identification of Max target genes, and this method can also be applied for other transcription factors.

Estrogen receptor beta in breast cancer.

Estrogen is essential for normal growth and differentiation in the mammary gland. It also supports growth of approximately 50% of primary breast cancers. For this reason, removal of estrogen or blocking of its action with the anti-estrogen, tamoxifen, is the main treatment for estrogen receptor alpha (ERalpha)-positive tumors. In 1996, when oncologists became aware of a second ER, ERbeta, there was some doubt as to whether this receptor would be of importance in breast cancer because the clinical consensus was that responsiveness to tamoxifen is related to the presence of ERalpha in breast cancer. Today we know that ERalpha and ERbeta have distinct cellular distributions, regulate separate sets of genes and can oppose each other's actions on some genes. We also know that ERbeta is widely expressed in both the normal and malignant breast and that there are proliferating cells in the breast which express ERbeta. In this review we summarize what is known about ERbeta in breast cancer and examine the possibility that ERbeta-selective ligands may well represent a useful class of pharmacological tools with a novel target, namely proliferating cells expressing ERbeta.

Estrogen receptors and proliferation markers in primary and recurrent breast cancer.

To elucidate the clinical importance of estrogen receptor (ER) beta in breast cancer, 29 archival primary breast cancer specimens, six locally recurrent cancers, and five benign mammary tumors were examined histochemically for ERalpha, ERbeta and the proliferation markers Ki67 and cyclin A. In benign tumors, most epithelial cells contained ERbeta, but ERalpha was rare. In primary cancers, both ERalpha and ERbeta occurred in epithelial cells, the presence of ERbeta being associated with elevated expression of Ki67 and cyclin A, and ERalpha with decreased levels. Thus, the highest content of proliferation markers was seen in primary cancers that were ERalpha(-) ERbeta(+). Most Ki67-containing cells coexpressed ERbeta, but few showed ERalpha. In locally recurring cancers, ERalpha, ERbeta, and Ki67 were more highly expressed than in the corresponding primary tumors, and many cells containing ERbeta, but few with ERalpha, expressed Ki67. Surprisingly, ERbeta, but not ERalpha, was seen in the stromal cells of both primary and recurrent cancers. Because the response of breast cancers to tamoxifen therapy is correlated with the presence of ERalpha, cancer cells that lack ERalpha but contain ERbeta and proliferation markers represent a novel population of apparently proliferating cells that probably are not targeted by the current antiestrogens. Thus, appropriate ERbeta-specific ligands, perhaps in combination with tamoxifen, may be useful in improving the treatment of breast cancers.

A 5-year follow-up study on the use of a levonorgestrel intrauterine system in women receiving hormone replacement therapy.

OBJECTIVE: To investigate endometrial histology, bleeding, and the effects of replacing the levonorgestrel intrauterine system (LNG IUS, Mirena/Levonova, Leiras Oy, Turku, Finland) after 5 years of combined use with estrogen. DESIGN: Prospective cohort study. SETTING: Private outpatient clinic. PATIENT(S): Forty postmenopausal women started hormone replacement therapy with LNG IUS and either a patch (50 microg/24 h) or oral (2 mg) estradiol valerate protocol. Thirty-nine completed 12 months of treatment. Twenty-nine of them had used LNG IUS with continuous estradiol replacement therapy for 5 years. Seven of them volunteered to a 3-month treatment-free period before reinsertion; 22 opted for immediate reinsertion. INTERVENTION(S): Endometrial sampling for histology, endometrial thickness, and location of the LNG IUS by ultrasound at removal and after washout. The women completed bleeding diaries from 3 months prior to removal to 3 months after reinsertion. MAIN OUTCOME MEASURE(S): Endometrial histology was evaluated and estrogen and progestin effects were also investigated. Endometrial thickness was measured. Bleeding was examined based on bleeding diaries. The investigator and the women evaluated the insertion, removal, and reinsertion of the LNG IUS. RESULT(S): At 6 and 12 months endometrial histology was nonproliferative. At removal, all endometria were suppressed and a strong progestin effect was seen. The thickest endometrium was 3.6 mm. After washout, all seven endometria were atrophic. Before the IUS was replaced, 26 women were amenorrheic, whereas three had minor spotting. After replacement 5 women had no bleeding and an additional 10 women had only spotting. The bleeding or spotting discontinued within 18 days. The insertion of LNG IUS was characterized as easy by the investigator and it was well tolerated by the women. Cervical dilatation and/or paracervical blockade was used in 10 insertions. CONCLUSION(S): Intrauterine levonorgestrel effectively protects against endometrial hyperplasia. In most women it induces amenorrhea, which is only temporarily affected by replacement of the LNG IUS.

The levels of trypsinogen isoenzymes in ovarian tumour cyst fluids are associated with promatrix metalloproteinase-9 but not promatrix metalloproteinase-2 activation.

Proteolysis mediated by matrix metalloproteinases (MMPs) and serine proteinases is associated with cancer invasion and metastasis. Activation of latent proMMPs, and especially the proforms of the type IV collagen degrading gelatinases A and B (proMMP-2 and proMMP-9), is thought to be a critical step in this process. We have recently found that human tumour-associated trypsin-2 is a potent activator of proMMP-9 and it also activates proMMP-2 in vitro. Trypsinogen, MMP-2, and MMP-9 are expressed in ovarian cancer. To elucidate the function of trypsin in vivo, we studied whether high concentrations of trypsinogen-1, trypsinogen-2, their alpha(1)-proteinase inhibitor (API) complexes, and tumour-associated trypsin inhibitor (TATI) are associated with proMMP-2 and proMMP-9 activation in ovarian tumour cyst fluids. Zymography and immunofluorometric analysis of 61 cyst fluids showed a significant association between high trypsin concentrations and the activation of MMP-9 (P = 0.003-0.05). In contrast, the trypsin concentrations were inversely associated with the activation of MMP-2 (P = 0.01-0.02). Immunohistochemical analysis of ovarian tumour tissue demonstrated expression of trypsinogen-2 and TATI in the secretory epithelium. MMP-2 was detected both in stromal and epithelial cells whereas MMP-9 was detected in neutrophils and macrophage-like cells in stromal and epithelial areas. These results suggest that trypsin may play a role in the regulation of the MMP-dependent proteolysis associated with invasion and metastasis of ovarian cancer.

Prospective study on gynaecological effects of two antioestrogens tamoxifen and toremifene in postmenopausal women.

To assess and compare the gynaecological consequences of the use of 2 antioestrogens we examined 167 postmenopausal breast cancer patients before and during the use of either tamoxifen (20 mg/day, n = 84) or toremifene (40 mg/day, n = 83) as an adjuvant treatment of stage II-III breast cancer. Detailed interview concerning menopausal symptoms, pelvic examination including transvaginal sonography (TVS) and collection of endometrial sample were performed at baseline and at 6, 12, 24 and 36 months of treatment. In a subgroup of 30 women (15 using tamoxifen and 15 toremifene) pulsatility index (PI) in an uterine artery was measured before and at 6 and 12 months of treatment. The mean (+/-SD) follow-up time was 2.3 +/- 0.8 years. 35% of the patients complained of vasomotor symptoms before the start of the trial. This rate increased to 60.0% during the first year of the trial, being similar among patients using tamoxifen (57.1%) and toremifene (62.7%). Vaginal dryness, which was present in 6.0% at baseline, increased during the use of tamoxifen (26.2%) and toremifene (24.1%). Endometrial thickness increased from baseline (3.9 +/- 2.7 mm) to 6.8 +/- 4.2 mm at 6 months (P< 0.001), and no difference emerged between the 2 regimens in this regard. Before the start of the antioestrogen regimen, the endometrium was atrophic in 71 (75.5%) and proliferative in 19 of 94 (20.2%) samples; 4 patients had benign endometrial polyps. During the use of antioestrogen altogether 339 endometrial samples were taken (159 in tamoxifen group, 180 in toremifene group). The endometrium was proliferative more often in the tamoxifen group (47.8%) than in the toremifene group (32.2%) (P< 0.0001). 20 patients had a total of 24 polyps (17 in tamoxifen and 9 in toremifene group, P< 0.05) during the use of antioestrogens. One patient in the toremifene group developed endometrial adenocarcinoma at 12 months, and one patient had breast cancer metastasis on the endometrium. Tamoxifen failed to affect the PI in the uterine artery, but toremifene reduced it by 15.0% (P< 0.05) by 12 months. In conclusion, tamoxifen and toremifene cause similarly vasomotor and vaginal symptoms. Neither regimen led to the development of premalignant endometrial changes. Our data suggest that so close endometrial surveillance as used in our study may not be mandatory during the first 3 years of use of antioestrogen treatment.

Chlamydia trachomatis seropositivity is associated both with stillbirth and preterm delivery.

The cause of stillbirth and preterm delivery is often unknown. We studied the prevalence of Chlamydia trachomatis antibodies in mothers with stillbirth and preterm labor. Serum specimens from 72 mothers with stillbirth after the 21st gestational week, and from 48 mothers with preterm delivery between gestational weeks 23 and 29, both from the greater Helsinki area, and cord blood from 96 consecutive liveborn deliveries at the Department of Obstetrics and Gynecology, the University of Helsinki, were studied for antibodies to C. trachomatis immunotypes CJHI, GFK and BED by microimmunofluorescence test. The prevalence of C. trachomatis antibodies was highest, 33.3%, in mothers with stillbirth, 18.8% in mothers with preterm delivery, and 10.4% in cord blood. The IgM seropositivity rate was high among mothers with preterm delivery (8.3%). We conclude that C. trachomatis IgG antibodies are frequently detected in sera from mothers with stillbirth, suggesting past infection, while mothers with preterm delivery often have serum IgM antibodies, suggesting of acute infection.

Reproductive features in women developing ovarian granulosa cell tumour at a fertile age.

Ovarian granulosa cell tumour (GCT) is a rare malignancy, which has been linked to both infertility and infertility treatment with ovulation inducers. The reproductive features were analysed of 146 women with GCT diagnosed between 1956 and 1996. During the study period no changes were found in the mean age (53 years), menopausal status (59% postmenopausal), parity (32% nulliparous) or tumour size or stage at diagnosis. The clinical features in women with GCT at fertile age were compared with GCT diagnosed later in life and to population-based data. Nulliparity (50%) and history of infertility (22%) were more frequent if the tumour occurred at fertile age (n = 50). Of the 12 infertile cases, seven had anovulatory infertility (58%); 11 occurred during the era of ovulation inducers, but only five had used these drugs (clomiphene citrate in five patients, gonadotrophins in two, and tamoxifen in one patient) and no patient had undergone in-vitro fertilization. Endometrial hyperplasia was associated with GCT at all ages, while endometrial cancer was found solely after the age of 45 years. In conclusion, GCT at fertile age is associated with nulliparity and with a clinical presentation of anovulatory infertility, while GCT later in life is associated with a more normal average fertility pattern and with occurrence of endometrial cancer.

Effect of an estradiol gel with monthly or quarterly progestogen on menopausal symptoms and bleeding.

OBJECTIVE: To assess the acceptability, efficacy and endometrial safety of transdermal estradiol gel (Divigel/Sandrena) combined with monthly or quarterly oral progestogen (medroxyprogesterone acetate). METHODS: This 12-month, multicenter, open-label study was carried out at 12 study centers in Finland and Sweden. A total of 395 postmenopausal women received 1 mg estradiol in 1 g gel, daily, with oral medroxyprogesterone acetate 10 mg for the first 12 days every month (groups I and III) or every 3 months (group II). The main outcome measures were relief of climacteric symptoms, bleeding patterns and endometrial safety. RESULTS: All regimens reduced the severity of hot flushes, sweating episodes and vaginal dryness. In groups I and III, approximately 80% and 70% of women, respectively, had regular monthly withdrawal bleeding (excepting the first cycle), with irregular bleeding in 8.3% and 5.3% of treatment months. In group II, approximately 94% of women had regular tri-monthly withdrawal bleeding, with irregular bleeding in 10.7% of the treatment months. Endometrial hyperplasia was observed in 0.3% of women. More than 87% of subjects completed the study, and 97% of these rated the gel as acceptable or convenient. CONCLUSIONS: Both the 1- and 3-month regimens were equally effective in controlling climacteric symptoms and protecting against endometrial hyperstimulation. The bleeding patterns were comparable between groups and were similar to those reported for oral estrogens. Estradiol gel was highly acceptable to the majority of women.

A novel method to culture laryngeal human papillomavirus-positive epithelial cells produces papilloma-type cytology on collagen rafts.

A novel method to culture human papillomavirus (HPV) positive laryngeal epithelial cells is described. Biopsies of laryngeal papillomas and of HPV-positive laryngeal mucosa were first cultured as a monolayer in which irradiated laryngeal fibroblasts originally derived from a papilloma (PPLF-XR) patient served as feeder cells. When these fourth or fifth passage epithelial cells were transferred to allow growth on an organotypic growth base (collagen raft containing unirradiated PPLF), they grew as a multilayer. This layer showed features typical of HPV infection with koilocytosis, parakeratosis, and isolated dyskeratotic cells. Based on in situ hybridisation, the original tumour sections and epithelial cells from each monolayer passage, as well as the collagen raft sections, contained HPV DNA. Our results show that HPV-infected epithelial cells can be maintained during passages in monolayer culture and that PPLF can support the growth of these cells well. The monolayer cell culture and the collagen raft, the latter providing differentiation-promoting effects, appears to facilitate maintenance of the infected cells and of the viral genome.

Controlled release gel of ibuprofen and lidocaine in epidural use--analgesia and systemic absorption in pigs.

PURPOSE: Reduction of the frequency of injections and localization of the absorption of drug molecules to the injection site would be of great advantage in epidural pain treatment. The epidural use of a controlled release gel of lidocaine and ibuprofen was studied. METHODS: The effect of a poloxamer gel (25%) containing 2% lidocaine.HCl and 2% ibuprofen.Na on the duration of analgesia after epidural administration to pigs was compared with drug in solution. Analgesia was assessed by observing the motor function and the nociceptive reflex-withdrawal response to painful pressure stimulation on the feet. Pharmacokinetic and histological examinations were performed. RESULTS: Analgesia lasted significantly longer after epidural lidocaine gel injection in comparison with the solution. The gel prolonged the systemic absorption, thereby increasing the epidural availability of lidocaine for spinal analgesia. Although the absorption of ibuprofen was prolonged after epidural gel injection, the duration of analgesia as compared with the solution was not prolonged. After epidural injection, only slight inflammatory changes were observed in the tissue structures of the epidural space, but none in the spinal cord. CONCLUSIONS: These results demonstrate poloxamer gel to be a promising controlled-release, injectable epidural formulation for the management of pain.

Distinct chromosomal imbalances in uterine serous and endometrioid carcinomas.

Endometrial carcinoma shows various histological types that differ in their clinical presentation and prognosis. Comparative genomic hybridization was used to detect gains and losses of DNA sequences along all chromosome arms in 24 uterine serous and 24 uterine endometrioid carcinomas. In serous carcinomas, extensive genetic aberrations were detected in 17 of the 24 specimens, with a mean of 5.7 changes per tumor. The most frequent gains occurred at 3q (50%), 8q (33%), 5p (29%), 6p (29%), and 1q (29%), and the most common losses were located at 4q (17%), 15q (17%), and 18q (17%). Tumors exhibiting DNA copy number changes were associated with shorter overall survival. In endometrioid carcinomas, genetic aberrations were less frequent and simpler than in serous carcinomas. DNA sequence copy number changes were observed in 12 of the 24 cases, with a mean of 1.5 changes per tumor. The most frequent aberrations were gains at 1q (29%), 2q (13%), and 8q (13%). Losses were rarely observed. The diverging pattern of genetic changes observed in uterine serous and endometrioid carcinomas suggests different pathways of carcinogenesis in these tumor types.

Effects of transdermal oestrogen therapy in postmenopausal women: a comparative study of an oestradiol gel and an oestradiol delivering patch.

OBJECTIVE: To compare the efficacy, safety and tolerability of an oestradiol gel (1.0 mg of oestradiol daily, Divigel/Sandrena) with those of an oestradiol delivering patch (delivering 50 micrograms oestradiol/24 h, Estraderm TTS) in hormone replacement therapy of postmenopausal women. Dydrogesterone tablets (Terolut), 10 mg daily for the first 12 days of every month, were used as the progestogen component of the therapy. MAIN OUTCOME MEASURES: The effect of treatment on clinical symptoms and on endometrium, total body bone mineral density and lipid metabolism as well as the tolerability of the treatments with special emphasis on skin irritation and compliance were evaluated. DESIGN: An open, randomised, controlled, parallel-group trial of 12 months' duration. SETTING: The Medical Clinic of Kalevankatu, Helsinki, Finland. PARTICIPANTS: One hundred twenty postmenopausal women were treated with transdermal oestradiol combined with dydrogesterone. In addition, 25 women without HRT served as a reference group for the bone mineral density measurements. RESULTS: Both treatment regimens were equally effective in alleviating climacteric symptoms, preserving bone mineral density and were equally safe. A trend towards heavier bleeding was detected in patients treated with the oestradiol delivering patch. A statistically nonsignificant decrease of total cholesterol and triglyceride concentrations but no change in high-density lipoprotein cholesterol concentration was observed in both groups. The acceptability of the treatment was higher in the gel group (96.4%) than in the patch group (90.7%). Only two (3.3%) women using the oestradiol gel complained of skin irritation whereas 28 patients (46.7%, P < 0.001) using the oestradiol delivering patch reported this adverse effect. CONCLUSIONS: Both the oestradiol gel and the oestradiol delivering patch are equally effective in hormone replacement therapy but the gel preparation is less irritative to the skin.

Malignant tumors of the ovary or the breast in association with infertility: a report of thirteen cases.

BACKGROUND: Many questions have been raised recently about the relationship between infertility, fertility drugs and cancer. This prompted us to evaluate our patients having ovarian or breast cancer with a known history of infertility. METHODS: We report thirteen women who had been examined and/or treated for infertility before the occurrence of malignant tumors of the ovary or the breast at an age under 50 years in 1990-1995 in our unit. RESULTS: Mean age of the patients was 35 years (s.d. 5.9 years, range 28-47 years). Of the 11 ovarian tumors, one was a malignant teratoma, two were granulosa cell tumors and eight epithelial ovarian cancers. Ten women had received either clomiphene citrate alone or together with gonadotrophins, one had used only gonadotrophins, and in two patients ovarian cancer was detected during an infertility work-up but before any treatment. Four women had used clomiphene for more than twelve cycles. Two patients had ductal breast cancer. CONCLUSIONS: Our patients emphasize the need for follow-up and long-term prospective studies in infertile women who have been evaluated or treated for infertility.

Complement-regulatory proteins in ovarian malignancies.

Ovarian cancer has features that makes it well-suited for MAb adjuvant immunotherapy. Several of the MAbs used in clinical trials mediate cancer cell destruction by activation of complement (C). In this study, therefore, we examined the ability of ovarian-tumor cells to resist C attack. We found that the C regulators membrane cofactor protein (MCP, CD46) and protectin (CD59) were strongly expressed in the tumor cells in all 28 benign and malignant tumors examined. Decay-accelerating factor (DAF; CD55) was more heterogeneously expressed, and only 75% of the tumors exhibited a moderate amount of DAF in the tumor cells. In adenoma cells, CD59 and DAF were preferentially located apically, while in adenocarcinoma cells they were expressed also at the basolateral cell surface. The ovarian-carcinoma cell lines SK-OV-3, Caov-3, SW626 and PA-1 expressed both the 58- and the 68-kDa isoforms of MCP. DAF was present as a glycosyl-phosphatidylinositol(GPI)-anchored 70-kDa glycoprotein. The surface-expression level of DAF varied, and correlated with the vulnerability of the cells to C-mediated lysis. CD59 was expressed as a GPI-linked 19- to 25-kDa protein exhibiting multiple glycosylation variants. The surface expression of CD59 correlated with the amount of the main 1.9 + 2.1-kb CD59 mRNA transcripts. Neutralization of CD59 with an anti-CD59 MAb significantly enhanced C-mediated killing of the cell lines. Low expression of C regulators on the PA-1 teratocarcinoma cell line was associated with high sensitivity to C lysis. Thus, the expression of C regulators on malignant ovarian cells may constitute a tumor escape mechanism, and is a critical parameter to be examined when MAb therapy is being considered.

Evidence for divergence of DNA copy number changes in serous, mucinous and endometrioid ovarian carcinomas.

Comparative genomic hybridization was applied to detect and map changes in DNA copy number in 24 well or moderately differentiated epithelial ovarian carcinomas (eight serous, eight mucinous and eight endometrioid carcinomas). Twenty-three of the 24 tumours showed changes in DNA copy number in one or several regions (median 4, range 1-17). Gains were more frequent than losses (ratio 1.6:1.0). The most frequent gains occurred in chromosomes 1q (38%), 2p (29%), 7q (25%), 8q(38%) and 17q (38%), and the most common losses were located in chromosomes 8p (21%), 9p (25%) and 13q (21%). High-level amplifications were detected in seven tumours at 1q22-32, 2p15-22, 3qcen-23, 6p21-22 and 8q. In the three histological subtypes the copy number karyotypes showed substantial differences. Gains at 1q were observed in endometrioid (five cases) and serous tumours (four cases). Increased copy number at 10q was seen in endometrioid tumours only (four cases), whereas gains at 11q occurred mostly in serous tumours (four cases). In mucinous tumours, the most common copy number change was a gain at 17q (six cases). The results show that, in epithelial ovarian carcinoma, changes in DNA copy number are a rule rather than an exception, chromosomes 1, 2, 7, 8, 9, 13 and 17 being the most frequently affected. The diverging pattern of genetic changes seen in epithelial ovarian carcinomas with different histological phenotypes suggests that various pathways may lead to tumorigenesis and/or progression in these subgroups.

Chlamydia trachomatis detected in human placenta.

AIMS: To evaluate the relation between Chlamydia trachomatis infection and stillbirth, placental tissue was studied for the presence of C trachomatis. METHODS: Paraffin wax embedded placental tissue of a stillbirth fetus, born at the 36th week of gestation to a 21 year old mother with high serum antibody titres to C trachomatis immunotypes during pregnancy and who was culture positive to C trachomatis three years previously, was studied by in situ hybridisation, polymerase chain reaction, and immunohistochemistry for the presence of C trachomatis. RESULTS: C trachomatis was detected in placental specimens by in situ hybridisation and alkaline phosphatase antialkaline phosphatase staining in several sections, whereas control tissues were uniformly negative, indicating the presence of C trachomatis nucleic acid and antigen in the placenta. CONCLUSION: This is the first reported case in which C trachomatis has been demonstrated in the human placenta.

Ezrin expression in stromal cells of capillary hemangioblastoma. An immunohistochemical survey of brain tumors.

Ezrin is a cytoskeleton-associated protein that appears to link actin filaments to the plasma membrane. Immunocytochemical studies suggest that ezrin is expressed in epithelial cells but not in mesenchymal cells. In addition, ezrin is expressed by certain epithelial tumors, such as renal cell adenocarcinomas. Ezrin serves as a tyrosine kinase substrate, and is phosphorylated in epidermal growth factor-stimulated cells. Ezrin may thus mediate regulatory signals in different cell functions. We studied the distribution of ezrin in 104 cases of primary tumors of the central nervous system (CNS) by immunocytochemistry. Special interest was focused on capillary hemangioblastoma, owing to its resemblance to renal cell adenocarcinoma, and on malignant gliomas, owing to their frequent epidermal growth factor receptor amplification. The stromal cells of hemangioblastomas were found to be strongly positive for ezrin. No expression was detected in gliomas and, except for hemangioblastomas, ezrin expression was restricted to those few CNS tumors that show epithelial differentiation, ie, choroid plexus papillomas, craniopharyngiomas, ependymomas, and cysts. The diffuse cytoplasmic expression of ezrin in the stromal cells of capillary hemangioblastoma may indicate that stromal cells overexpress ezrin or express ezrin with deficient binding properties.

Controlled release of lidocaine from injectable gels and efficacy in rat sciatic nerve block.

PURPOSE: Methods of delaying the action of local anesthetics are important, since short duration of action limits their use in the treatment of postoperative and chronic pain. The present study evaluated the use of low-viscosity gels in prolonging the release of lidocaine. METHODS: Release of lidocaine from 2% lidocaine.HCl containing methylcellulose (MC), hydroxypropylmethylcellulose (HPMC), sodiumcarboxymethyl cellulose (CMC), and poloxamer 407 (PO) gels was studied in phosphate buffer, pH 7.4, at 37 degrees C. Commercial metylcellulose gel (MCcom) served as control. The in vivo efficacy of the respective gel formulations were evaluated in rats. The gel was injected into the vicinity of the sciatic nerve and nociception and motor function were tested. RESULTS: The cumulative amount of lidocaine released during 8 hr was slowest from the PO gel, followed by the CMC, HPMC and MC gels. The antinociceptive effect was not prevented by the motor block and lasted longest with the PO gel. Good linear and rank order correlation was obtained between in vitro and in vivo results. The microscopic examination of the tissue samples revealed only mild or no irritation of the skeletal muscle tissue by the PO, HPMC, and CMC gels. CONCLUSIONS: Based on these results poloxamer gel proved to be the most promising carrier for lidocaine.

Altered growth behavior of human cervical epithelial cells transfected by HPV type 16 and 18 DNA.

Transfection of primary human cervical epithelial (HCE) cells with full-length HPV type 16 and 18 DNAs resulted in cell lines that could grow continuously. Four HPV DNA-immortalized cell lines were established. Morphologically the immortalized cells resembled primary HCE cells. Electron microscopy showed that they contained desmosomes and keratin filaments, which are characteristic structural components of epithelial cells. Each cell line had a unique integration pattern of HPV DNA but the transcription patterns were similar in the 3 HPV 16 DNA-immortalized cell lines. The expression patterns of viral DNA in the HPV 18 DNA-immortalized cell line were similar to that in HeLa cells, suggesting transcription of mainly early viral genes. The cell lines, unlike HeLa and SiHa carcinoma cells, did not form tumors in nude mice or grow in soft agarose, but collagen raft culture indicated that the immortalized cell lines had lost the capacity of normal differentiation compared with primary HCE cells. Morphologically, the aberrant differentiation of the immortalized cells showed great resemblance to cervical intra-epithelial neoplasia. The altered pattern of growth and differentiation of human cervical epithelial cells transfected by HPV 16 and 18 DNAs is in agreement with the view that HPV types 16 and 18 play an important role at an initial step of human cervical epithelial carcinogenesis but that co-carcinogenic factors are necessary for full malignant transformation in vivo.