Involvement of thyrotropin-releasing hormone receptor, somatostatin receptor subtype 2 and corticotropin-releasing hormone receptor type 1 in the control of chicken thyrotropin secretion.

Thyrotropin or thyroid-stimulating hormone (TSH) secretion in the chicken is controlled by several hypothalamic hormones. It is stimulated by thyrotropin-releasing hormone (TRH) and corticotropin-releasing hormone (CRH), whereas somatostatin (SRIH) exerts an inhibitory effect. In order to determine the mechanism by which these hypothalamic hormones modulate chicken TSH release, we examined the cellular localization of TRH receptors (TRH-R), CRH receptors type 1 (CRH-R1) and somatostatin subtype 2 receptors (SSTR2) in the chicken pars distalis by in situ hybridization (ISH), combined with immunological staining of thyrotropes. We show that thyrotropes express TRH-Rs and SSTR2s, allowing a direct action of TRH and SRIH at the level of the thyrotropes. CRH-R1 expression is virtually confined to corticotropes, suggesting that CRH-induced adrenocorticotropin release is the result of a direct stimulation of corticotropes, whereas CRH-stimulated TSH release is not directly mediated by the known chicken CRH-R1. Possibly CRH-induced TSH secretion is mediated by a yet unknown type of CRH-R in the chicken. Alternatively, a pro-opiomelanocortin (POMC)-derived peptide, secreted by the corticotropes following CRH stimulation, could act as an activator of TSH secretion in a paracrine way.

Development of squamous cell carcinoma by two high-risk human papillomaviruses (HPVs), a novel HPV-67 and HPV-31 from bowenoid papulosis.

We report a patient with bowenoid papulosis (BP) involving two high-risk human papillomaviruses (HPVs) and the development of invasive squamous cell carcinoma (SCC). Our patient showed verrucous lesions on the penis, perianal area and groin that had been noted over the previous 8 years and had recurred after all therapeutic approaches. The perianal and left inguinal lesions revealed invasive SCC on histology. HPV-31 and HPV-67 sequences were detected by polymerase chain reaction from BP lesions of the perianal area and the shaft of the penis. HPV-31 has already been reported in BP as a high-risk HPV for the development of SCC, but HPV-67 is a novel one that has never been reported in BP. As HPV-67 has sequence homology to HPV-52 and HPV-58, it belongs to the family of HPV-16, a high-risk HPV group. Thus our patient showed two high-risk HPVs, i.e. HPV-31 and the novel HPV-67, which may be directly involved in the development of SCC.

Chicken lutropin acts like follitropin in rat ovarian follitropin receptor: an isoelectric focusing study.

This study investigates whether chicken lutropin (LH) specifically binds to rat ovarian follitropin (FSH) receptor and exerts FSH-like bioactivity. Glycoprotein fraction, prepared from the chicken anterior pituitary gland, was fractionated using isoelectric focusing within a pH range of 3.5-11. Analysis of the focused fractions, by a radioreceptor assay (RRA) specific for FSH in rats using rat ovarian homogenate as receptor source, and 125I-labeled rat FSH as radioligand, detected a large component having an isoelectric point of 10.25. This focusing profile obtained by RRA was quite similar to that obtained by a specific radioimmunoassay (RIA) for chicken LH, but clearly different from that obtained by a specific RIA for chicken FSH, indicating this RRA specifically recognizes chicken LH. Chicken LH fraction prepared from the electrofocused material was used for further studies. The chicken LH preparation was three times more potent than rat FSH in the RRA in displacing the radioligand bound to rat ovarian receptor, while chicken LH facilitated an 8-fold less production of estradiol in dispersed rat granulosa cells than rat FSH. These results suggest that chicken LH acts like rat FSH in rat ovarian FSH receptor, but receptor-binding activity is much higher than biological activity.

Metastatic cervical carcinoma mimicking kidney abscess.

A 26-year-old female, who had been treated for cervical carcinoma, presented with high fever and right flank pain. A right renal abscess was initially suspected from the clinical symptoms and diagnostic imaging. However, pathologic findings for the right kidney revealed squamous cell carcinoma, which was consistent in type with the original cervical carcinoma. Demonstration of human papillomavirus 16 in tissues from both the renal tumor and the cervical carcinoma confirmed that the right kidney carcinoma was a metastasis from the cervical carcinoma. The role of interleukin-6 in occurrence of the unexplained fever is discussed.

Evaluation of a sensitive, in vitro bioassay for chicken thyroid stimulating hormone using FRTL-5 cells, a rat thyroid cell line.

An in vitro bioassay for mammalian thyroid stimulating hormone (TSH) based on TSH-induced cyclic adenosine 3',5'-monophosphate (cAMP) production in FRTL-5 cells, a rat thyroid cell line, was used to measure chicken TSH. The addition of chicken pituitary homogenate equivalent to > or = 25% of a chicken pituitary gland to cultured FRTL-5 cells increased cAMP within these cells in a dose-dependent manner. The glycoprotein fraction derived from the pituitary homogenate was further fractionated by isoelectric focusing within a pH range of 5 to 11. Analysis of the focused fractions by the bioassay detected three major components with isoelectric points of 9.30, 7.12, and 3.82, in addition to several minor ones distributed over a wide range of pH, from alkaline to acidic. The isoelectric focusing profile obtained by the bioassay was clearly different from those obtained by radioimmunoassay for chicken LH and radioreceptor assay for chicken FSH, indicating that fractions contained chicken TSH. The homogenate of the cephalic portion of the chicken anterior pituitary gland was 4.46 times more active than that of the caudal portion in the bioassay, which is consistent with previous findings on localization of TSH in the chicken pituitary. We conclude that the bioassay using FRTL-5 rat thyroid cells is a sensitive, specific, and time-saving method of measuring chicken TSH.

Human papillomavirus DNA in urine specimens of men with condyloma acuminatum.

BACKGROUND AND OBJECTIVES: Because warts are often found in the male urethra, human papillomavirus (HPV) may well be present in urine of patients with urethral condylomata. GOAL: To detect HPV DNA in urine specimens of men with condylomata acuminata using polymerase chain reaction. STUDY DESIGN: Forty-seven urine specimens and 25 paraffin-embedded tissues of condylomata acuminata were obtained from men. Of the 47 urine specimens, 29 were from patients with urethral condylomata, 3 from patients with penile condylomata only, and 15 from control subjects without condylomata. Both L1 consensus primers and type-specific primers for-HPV 6, 11, 16, 18, and 33 were used. RESULTS: HPV DNA was detected in 22 of the 29 (76%) urine specimens from patients with urethral condylomata, in none of the 3 urine specimens from patients with penile condylomata, and in none of the 15 controls. Paraffin-embedded tissues of all 25 condylomas were positive for HPV DNA. The HPV types detected in urine were identical to those detected in urethral condylomas. CONCLUSIONS: HPV DNA is present in urine of patients with urethral condylomata. Urine may be used for noninvasive screening of asymptomatic HPV infections of the male genital tract. Detection of HPV DNA in urine may be useful for monitoring the response to treatment of urethral condylomata.

Human papillomavirus in squamous cell carcinoma of the vulva by polymerase chain reaction.

OBJECTIVE: To investigate the prevalence of human papillomavirus (HPV) DNA in squamous cell carcinoma of the vulva by polymerase chain reaction (PCR). METHODS: Archival diagnostic phase biopsies from 74 patients with squamous cell carcinoma of the vulva were investigated for HPV DNA by PCR. We used both consensus primers located in the open reading frame L1 and type-specific primers for HPV 6 (open reading frame E5), HPV 11 (open reading frame L1), HPV 16, HPV 18, and HPV 33 (open reading frame E6). RESULTS: HPV DNA was detected in 27 (36%) of the 74 patients, of whom 19 had HPV 16, nine had HPV 18, one had HPV 33, and one had unclassified HPV DNA. No case of HPV type 6 or 11 was detected. Two squamous cell carcinomas were positive for both HPV 16 and 18, and one was positive for both HPV types 16 and 33. Three squamous cell carcinomas positive for E6 gene using type-specific primers were negative using L1 consensus primers. CONCLUSION: Our PCR methods using both consensus open reading frame L1-derived primers and type-specific open reading frame E6-derived primers of HPV types 16, 18, and 33 seemed to be an appropriate combination for the detection of HPV DNA in archival tissues of vulvar carcinoma. Both HPV types 16 and 18 were associated with squamous cell carcinoma of the vulva, although the prevalence of HPV 16 was considerably lower than in cervical carcinoma. It appears that vulvar and cervical carcinomas are not identical etiologically and that factors other than HPV are important in vulvar carcinogenesis.

Human papillomavirus DNA in uterine cervix squamous cell carcinoma and adenocarcinoma detected by polymerase chain reaction.

BACKGROUND: Substantial clinical, epidemiologic, and experimental evidence has reinforced the role of high risk human papillomavirus (HPV) types in the development of cervical carcinoma. The authors investigated HPV in the uterine cervix squamous cell carcinomas and adenocarcinomas of Finnish patients. METHODS: Specimens from 352 patients with uterine cervix squamous cell carcinomas and 108 with adenocarcinoma were examined for HPV DNA by polymerase chain reaction. The authors used consensus primers located in the L1 region, as well as HPV16, 18, and 33 type-specific primers located in the E6 region. RESULTS: HPV DNA was detected in 324 of 352 squamous cell carcinomas (92%), and 81 of 108 adenocarcinomas (75%). Two-hundred seventy-four of 352 squamous cell carcinomas (78%) and 18 of 108 adenocarcinomas (17%) contained HPV16 DNA, whereas 55 of 352 squamous cell carcinomas (16%) and 60 of 108 adenocarcinomas (56%) contained HPV18 DNA. Eight squamous cell carcinomas and 4 adenocarcinomas were positive for HPV33. Twenty-eight squamous cell carcinomas and 5 adenocarcinomas were positive for either HPV16 and HPV18 or HPV16 and HPV33. Unclassified HPV DNA was detected in 17 squamous cell carcinomas and 4 adenocarcinomas. Twenty-eight squamous cell carcinomas and 9 adenocarcinomas, which were positive for E6 DNA using type-specific primers, were negative for the L1 gene. All 460 cervical specimens were tested twice with identical results. CONCLUSIONS: HPV DNA was highly prevalent in both uterine cervix squamous cell carcinoma and adenocarcinoma. HPV16 was detected more often in squamous cell carcinoma and HPV18 was detected more often in adenocarcinoma. Both consensus structural L1 gene-derived primers and type-specific viral E6 oncogene-derived primers were necessary to detect HPV DNA in cervical carcinoma.

Time-resolved fluoroimmunoassay (TR-FIA) of gonadotropins.

Time-resolved fluoroimmunoassay (TR-FIA), which has recently been developed as a non-isotopic immunoassay, was intended for assay of gonadotropins (LH and FSH). Ovine LH or FSH was labeled with N1-(p-isothiocyanatobenzyl)-diethylenetriamine-N1, N2, N3, N3-tetraacetic acid Eu-chelate (DTTA), and competitive binding assay was performed using a 96-weel titer plate previously coated with a second antibody, followed by measurement using time-resolved fluorometry. TR-FIA for standard ovine LH (NIDDK, LH-I-3) or FSH (NIDDK, FSH-I-1) had a sensitivity of about 25 pg/50 microliters sample. The assay system was applied to heterologous assay of porcine gonadotropins. Linearity was obtained by the dilution test using medium from primary culture of porcine anterior pituitary cells. Intra- and inter-assay coefficients of variation of LH and FSH determination in 31 different porcine samples were satisfactorily low, between 3.5 and 8.1% (intra-assay) and between 1.7 and 13.1% (inter-assay). Correlation coefficients between radioimmunoassay and TR-FIA were calculated to be 0.945 for LH and 0.978 for FSH. Stimulation of LH and FSH release with GnRH was observed by TR-FIA. This non-isotopic TR-FIA thus provides as good sensitivity, reproducibility and accuracy as conventional RIAs.

Monoclonal antibodies as probes to detect conformational changes in the rat cysteine proteinase inhibitor cystatin A.

Five monoclonal antibodies (MAbs), 77, 114, 138, 175 and 187, were established for rat cystatin A. MAbs 77, 114, 138 and 175 were shown to belong to the IgG1 subclass, whereas MAb 187 was an IgM. These MAbs partially suppressed inhibitory activity of rat cystatin A to papain. Their epitopes were mapped in detail on the molecule by examining the reactivities of the MAbs with NH2-terminally truncated forms and fragments of rat cystatin A by an enzyme-linked immunosorbent assay (ELISA), and by reactivity with the inhibitor on immunoblotting. In competitive binding assays the MAbs did not compete with each other, indicating that the epitopes recognized by these MAbs were substantially different. The conformational epitope recognized by the three MAbs 114, 138 and 175 belonged to one group that was highly sensitive to denaturation, but those epitopes were unchanged by NH2-terminal truncation. MAb 187 was able to recognize a linear epitope present in amino acid residues 15-50 in the NH2-terminal region. MAbs 77 and 114 reacted weakly with mouse cystatin A but not at all with human cystatin A, whereas MAb 187 reacted similarly with mouse cystatin A but at about half that level with human. The MAbs produced in this study should be useful tools for detecting conformational changes in the rat cystatin A molecule.

Detection of human papillomavirus deoxyribonucleic acid in penile carcinoma by polymerase chain reaction and in situ hybridization.

We investigated the prevalence of human papillomavirus types 16, 18 and 33 deoxyribonucleic acid (DNA) by polymerase chain reaction and the localization of human papillomavirus DNA by in situ hybridization using formalin-fixed, paraffin-embedded tissue specimens of penile carcinoma in Japan. Of 111 untreated penile carcinomas 70 (63.1%) were positive for human papillomavirus DNA by the polymerase chain reaction method. Human papillomavirus type 16 was identified in 68 penile carcinoma cases and type 18 in 2, whereas type 33 could not be detected. Of 12 treated penile carcinomas 2 (16.7%) were positive for human papillomavirus type 16. These data indicate that human papillomavirus type 16 DNA is the type most commonly associated with penile carcinoma. Human papillomavirus type 16 was also detected in lymph node metastasis of penile carcinoma, and that was the same as the human papillomavirus DNA type in the primary carcinoma. The in situ hybridization analysis found the human papillomavirus DNA to be localized in the nuclei of carcinoma cells.

[Detection of human papillomavirus DNA in cases of sexually transmitted diseases (STD)].

Human papillomavirus (HPV) has been detected on the genitalia without any macroscopic abnormality and the possibility of latent infection of HPV has been suggested. Using Vira Type (Toure Co.), we have detected 7 genotypes of HPV DNA under a high stringent condition on the genitalia of patients with sexually transmitted diseases (STD), who were suspected of having had many sexual partners. In male cases of STDs other than condyloma acuminatum, the HPV-positive rate of the glans and sulcus coronarius was 4.7% (5/106). In healthy men, the HPV-positive rate was 6.1% (2/33), while in chronic prostatitis cases it was 3.4% (7/205) and in benign prostatic hypertrophy cases HPV was not detected. In female cases of STDs other than condyloma acuminatum, the HPV-positive rate of uterine cervix was 5.1% (3/58). In pregnant women, the HPV-positive rate was 4.6% (9/197). With regard to the HPV-positive rate within different age groups of STD and non STD cases, the rate tended to be higher in young people. After several weeks, follow-up studies were conducted on HPV-positive cases. HPV DNA was detected in one case of 10 males STD cases and two of 10 pregnant women, and the HPV DNA was the same type as at the first examination. However, after 3-4 months, all three of these cases had become negative for HPV DNA.

Presence of human papillomavirus 6/11 DNA in condyloma acuminatum of the urinary bladder.

A Japanese woman with condyloma acuminatum of the urinary bladder is presented. The condyloma acuminatum lesion was resected endoscopically and human papillomavirus 6/11 DNA was detected. After treatment, there has been no recurrence of the disease.

[Immunosuppressive acidic protein (IAP) and immunosuppressive substance (ISS) in patients with renal cell carcinoma].

To clarify their usefulness as markers for renal cell carcinoma, serum immunosuppressive acidic protein (IAP) and serum immunosuppressive substance (ISS) were evaluated by TIA (turbidometric immunoassay) for IAP and by SRID (single radial immunodiffusion) for ISS. The mean level of IAP and ISS was beyond each upper normal limit (500, 700 micrograms/ml) in every stage, and especially high in the M1 group. The levels of IAP and ISS were significantly correlated with each other. The determination of IAP and ISS levels after treatment showed a good correlation to the clinical course of the disease. The positive rates of IAP and ISS increased as the stages progressed, respectively. When the influences of pretreatment IAP and ISS level on survival period were investigated, the low IAP or ISS level group (less than two times of the upper normal limit) tended to have a better prognosis than the high level group (more than two times of the upper normal limit) in the M1 patients. These findings suggested that IAP and ISS could be used as markers for monitoring a disease and predicting the prognosis in patients with renal cell carcinoma. As for the positive rate in the combination assay for IAP, TPA and ferritin, or ISS, TPA and ferritin, more than 80% of the patients with low stage renal cell carcinoma had at least one positive marker. This suggested that the combination assay of these three markers was clinically valuable as a disease monitor in patients with renal cell carcinoma.

Transitional cell carcinoma of the bladder or renal pelvis in children.

Transitional cell carcinoma (TCC) of the bladder or renal pelvis is rare in children. We report 2 children with TCC of the bladder and 1 with TCC of the renal pelvis. One of the two children with bladder carcinoma experienced frequent intravesical recurrences, which is in contrast to the usual clinical course of bladder carcinoma in children. In the 3rd child, renal pelvic carcinoma was found incidentally in a renal pelvis specimen removed during pyeloplasty.

[Studies of human papillomavirus (HPV) in urological tumors].

Urological tumors were examined for the presence of human papillomavirus (HPV) DNA by using Southern blot hybridization. In 20 male patients with condyloma acuminatum, HPV type 6 was found at 85% (17/20), HPV type 11 at 95% (19/20), HPV type 16 at 5% (1/20) and HPV type 18 at 0% (0/20). In 2 female patients with condyloma acuminatum, HPV types 6, 11, 16 and 18 were found at 100% (2/2), 100% (2/2), 50% (1/2) and 0% (0/2), respectively. All 6 of the patients who were positive for HPV type 6, were also positive for HPV type 11. Two patients were positive for HPV types 6, 11 and 16, the last of which was frequently found in penile cancer and uterine cervical cancer. In 6 patients with penile cancer, two patients were positive for HPV type 16 and negative for HPV types 6, 11 and 18. The remaining 4 patients were negative for all these HPV types. One patient who was positive for HPV type 16 had penile cancer after three previous episodes of penile condyloma acuminatum. From this information, a malignant change in the condyloma acuminatum was assumed to indicate the possible association of HPV type 16 with the process of malignant degeneration. HPV types, 6, 11, 16 and 18 were not detected in a female patient with vulvar cancer. Although HPV was thought to participate in the development of urological tumors except for external genital tumors, all patients examined, consisting of 2 with benign prostatic hypertrophy, 5 with prostatic cancer and 24 with bladder cancer, were negative for HPV types 6, 11, 16 and 18. Eight patients with bladder cancer were negative for HPV type 33.

[Detection of human papillomavirus DNA in the normal genitalia of men].

The study of human papillomavirus (HPV) in the genitalia of all normal males is difficult, therefore cases of chronic prostatitis, a common disease treated in urological clinics, were chosen to identify HPV DNA by using Vira Type (Life Technologies Inc.). Smears of glans and sulcus coronalius of 177 subjects, showed a HPV positive rate of 3.4%, while 86 cases of those cases were negative for HPV in urethral smears. The lack of clinical findings suggests that HPV is an asymptomatic infection. In a follow up examination of 5 HPV positive cases some weeks later, smears of glans, sulcus coronalius and urethra were all negative for HPV. Examination by Vira Type showed that HPV disappeared spontaneously in these cases.