Biomedical subjects
Publications and source records attributed to T Tohma.
Pit-1 is a pituitary-specific transcription factor that binds to and transactivates promoters of growth hormone- and prolactin-encoding genes. A chromosomal gene related to human Pit-1 isolated from human gene libraries was over 14 kb long and split into six exons. All of the splice donor and acceptor sites conformed to the GT/AG rule. The gene was mapped to human chromosome region 3p11.
We previously constructed a chromosome 2q-specific genomic library and isolated a number of microclones. In the present study, we first analyzed with Southern hybridization whether any of the microclones represent restriction fragment length polymorphisms (RFLPs), and then tried to map RFLP markers physically, using the recently developed chromosome microdissection/enzymatic amplification method. Of 13 clones analyzed, two were RFLP markers; a clone, pM2C83, showed a four-allele MspI RFLP, and the other, pM2C8, a two-allele RsaI RFLP. In order to assign the two polymorphic markers, two chromosomal segments, 2q32-q35 and 2q35-qter, on the chromosome 2 from a karyotypically normal person were microdissected, and the DNA from each segment was amplified with the polymerase chain reaction (PCR) using marker sequence-specific primers. With this method, both of the clones were assigned to 2q35. These two RFLP markers must be useful for linkage analysis of genetic diseases whose loci are at around 2q35.
Human chromosomal regions 8q23.3-q24.11 and 2q33-qter were microdissected, DNAs from the regions were amplified with the primer-linker method of polymerase chain reaction (PCR), and their DNA libraries were constructed by cloning into pUC19. The primer-linker PCR involved Sau3AI digestion of microdissected chromosomal DNAs, ligation of the digests to a 10mer DNA linker and 24mer primer, filling the recessed 3' ends, and PCR amplification using the 24mer DNA as a primer. A total of 3.5 x 10(4) pUC19 recombinants (8q library) from the 8q region and 5.0 x 10(4) pUC clones (2q library) from the 2q region were obtained. From the 8q library, 60 pUC clones were selected, while 88 pUC-clones were selected from the 2q library. These clones were Southern blot analyzed on hybrid cell panels with or without human chromosome 8 or 2. Twelve (20%) of the 60 8q-derived clones were unique DNA sequences, and 9 were subjected to deletion analysis in the genomic DNA of two patients, one with trichorhino-phalangeal syndrome (TRPS) type I and the other with TRPS type II, both with del(8) (q23.3q24.13). Five of the 9 pUC clones tested showed a one-copy density in both patients, an indication that the clones map to the region deleted in both patients. Screening a genomic DNA library constructed in the phage revealed a clone with a 9.4-kb insert and a one-copy density in both patients. From the 2q library, 15 (17%) of the 88 pUC clones obtained were unique sequences. When a phage library was screened, 8 clones were obtained: 4 were identical and 2 were overlapping sequences.(ABSTRACT TRUNCATED AT 250 WORDS)
We described cloning and characterization of an inversion breakpoint of chromosome 2 inv(2)(q35q37.3) observed in a patient with Waardenburg syndrome type I (WSI). Genomic cosmid clones containing the HuP2 gene, which was considered as a candidate for WSI, were isolated from a library constructed from the patient DNA. One of the clones contained the inversion breakpoint and revealed signals at both 2q35 and 2q37 by fluorescent in situ hybridization (FISH), indicating disruption of the HuP2 gene by the inversion. Our result further supports that the HuP2 gene is a candidate for Waardenburg syndrome type I and is located at q35.
An alternative method for amplification of DNA with unknown sequences was developed. This involves the direct ligation of a primer oligodeoxyribonucleotide itself to restricted DNA fragments with unknown sequences to be amplified by PCR. The oligonucleotide need not be phosphorylated and need not be annealed with its complementary oligonucleotide in advance for ligation. The ligation reaction seems to be independent of the concentration of unknown DNA, proceeds in short time, and is efficient. The ligation efficiency was more than 30% at a low concentration, 10 fg/microliters, of DNA. This method was applied to a microdissection/microcloning of the short arm of human chromosome 2. Of 65 clones screened for the highly repetitive sequences with total human genomic DNA, eleven (17%) were positive. Their inserts ranged in size from 150 to 1,200 bp (average, 460 bp). In Southern blot analysis, thirty consecutive clones all detected signals common to both total human genomic DNA and mouse-human hybrid cell DNA containing only chromosome 2 of human origin. Among them, 24 (80%) were unique sequences, and 6 (20%) were multi-copy (or intermediate-repeat) sequences. Thus, this method is simple and efficient, and provides an alternative way to amplify unknown DNA.
Clinical courses of 67 patients with carcinoma in situ (CIS) of the urinary bladder during 14 years from 1971 to 1984 were investigated according to the clinical type of CIS and treatment methods. CIS was classified into four types: the primary group included 18 patients who had neither prior nor simultaneous tumors of the urinary tract; the secondary group included 10 patients who had CIS diagnosed subsequent to the treatment of superficial papillary bladder tumor; the concurrent group included 14 patients who had CIS concomitantly with superficial papillary bladder tumor; and the nonpapillary T1 group included 25 patients who presented with CIS with concomitant nonpapillary T1 tumor. As a rule, the initial treatment was conservative (transurethral resection [TUR] or intravesical chemotherapy) for the primary, secondary, and concurrent CIS groups, whereas treatment was radical (total cystectomy or irradiation) for the nonpapillary T1 group. Five-year survival rates of the primary, secondary, concurrent, and nonpapillary T1 groups were 41%, 100%, 49%, and 68%, respectively. Secondary CIS revealed a rather good prognosis, probably due to the early detection of CIS and early application of intravesical chemotherapy when compared to other groups. Except for patients with nonpapillary T1 tumors, the 5-year rate of malignant progression (invasion or metastasis) and multiple recurrences leading to delayed cystectomy was 81% in 16 patients treated by TUR, whereas it was 39% in 21 patients treated by instillation therapy. It appears likely that intravesical chemotherapy was preferable to other conservative therapies as an initial treatment of CIS. Radical therapy, however, may be the choice for CIS with nonpapillary T1 tumors, ab initio.
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An in vivo staining test with 0.2 per cent methylene blue was applied to 129 patients with bladder tumor and 16 patients with chronic cystitis within a 6-year interval. Although normal mucosa did not pick up the stain nonpapillary in situ and microinvasive carcinomas did so frequently. Moderate dysplasia was stained in about half of the patients. The intensity of the stain in papillary tumors was correlated with the histologic anaplasia (grade). Grade 1 tumors were stained poorly or unstained in 86 per cent of the tests, whereas grades 2 and 3 tumors picked up the stain in 74 and 96 per cent of the tests, respectively. Even a tiny tumor, if poorly differentiated, was identified easily by the blue stain. The histologic anaplasia of tumors could be assessed roughly according to the intensity of the stain. However, chronic cystitis occasionally took up the stain, especially in cases of marked inflammatory infiltrate a deep stain was recognized. To differentiate nonpapillary early cancer from chronic cystitis the addition of a cytologic examination may be necessary.
Of 487 patients with urothelial bladder cancer seen during the past 22 years, 37 (7.6%) had other primary malignant tumors. In recent years the number of patients with double cancer markedly increased. The male/female ratio was 2.7:1 and the mean age at the time of diagnosis of the second primary cancer was 70 (56-84) years. Of the other organ cancers, 18 (47%) were in the digestive tract and 7 (18%) in the respiratory tract. Six patients had lesions that were diagnosed simultaneously and 31 had lesions that were diagnosed at intervals. The interval in 31 patients with metachronous double cancer ranged from 1 year and 1 month to 40 years; the mean interval was 9 years and 7 months.
From the studies on the stability of cefmetazole and cefazolin in feces of animals and children, and the determination of the fecal concentration of cefmetazole in three pediatric patients, the following results were obtained. 1) When cefmetazole was kept at 37 degrees C for 24 approximately 48 hours in feces of rats, rabbits, dogs and children, it showed no decrease of the potency. On the other hand, cefazolin kept in the same condition as that of cefmetazole showed a remarkable decrease of the potency. 2) Fecal excretions of cefmetazole were 0, 0.22 and 0.65% of total doses, respectively, in two pediatric patients of scarlet fever and one pediatric patient of serous meningitis.