Solid papillary carcinoma of the breast: report of two cases.

Two cases of solid papillary carcinoma of the breast were pathologically studied. Subjects included two female patients: a 76-year-old (Case 1) and a 43-year-old (Case 2). Both cases showed solid and papillary proliferation of spindle cells in expanded ducts, perivascular pseudorosettes, eosinophilic fine granules in an abundant cytoplasm, intracellular mucin production, and positivity for chromogranin A. Case 1 showed an aggregation form involving multiple ducts, admixture of signet-ring like cells, mucin pools, and small and large electron-dense granules and flocculent granules in electron microscopy. Case 2 showed a solitary and compact form in a dilated duct and an interlacing bundle pattern.

Use of lycorine and DAPI staining in Saccharomyces cerevisiae to differentiate between rho0 and rho- cells in a cce1/delta cce1 nuclear background.

In the yeast Saccharomyces cerevisiae, mutants are viable with large deletions (rho-), or even complete loss of the mitochondrial genome (rho0). One class of rho- mutants, which is called hypersuppressive, is characterised by a high transmission of the mutated mitochondrial genome to the diploid progeny when mated to a wild-type (rho+) haploid. The nuclear gene CCE1 encodes a cruciform cutting endonuclease, which is located in the mitochondrion and is responsible for the highly biased transmission of the hypersuppressive rho- genome. CCE1 is a Holliday junction specific endonuclease that resolves recombination intermediates in mitochondrial DNA. The cleavage activity shows a strong preference for cutting after a 5'-CT dinucleotide. In the absence of the CCE1 gene product, the mitochondrial genomes remain interconnected and have difficulty segregating to the daughter cells. As a consequence, there is an increase in the fraction of daughter cells that are rho0. In this paper we demonstrate the usefulness of lycorine, together with staining by 4',6-diamidino-2-phenylindole (DAPI), to assay for the mitotic stability of a variety of mitochondrial genomes. We have found that rho+ and rho- strains that contain CT sequences produce a large fraction of rho0 progeny in the absence of CCE1 activity. Only those rho- mitochondrial genomes lacking the CT recognition sequence are unaffected by the cce1 allele.

[Complete remission in a case of sigmoid colon cancer with multiple liver metastases-treatment with arterial chemotherapy and percutaneous microwave coagulation therapy].

A 59-year-old man was admitted to our hospital for advanced sigmoid colon carcinoma with synchronous multiple liver metastases. The patient received sigmoidectomy with regional lymph node dissection on June 8, 1998. We started intra-arterial combination chemotherapy on July 1, 1998. MMC (4 mg/body) was administered via rapid intra-arterial infusion on day 1. After MMC administration, 5-day intra-arterial continuous infusion of 5-FU at 500 mg/body/day was performed with oral administration of LV (30 mg/body/day). The treatment cycle was defined as every three weeks. The patient was treated with 4 courses of chemotherapy. From September 30, he received intra-arterial infusion of bolus MMC 4 mg/body, LV 6 mg/body and 5-FU 1,000 mg/body/4 hrs every two weeks with oral administration of Tegafur-uracil 400 mg/day. After 4 intra-arterial chemotherapy sessions, the metastatic liver tumors disappeared except for a focus in the right lobe. Therefore we decided to give the remnant liver metastasis percutaneous microwave coagulation therapy (PMCT). He obtained a complete remission in the liver metastases after two PMCT (70 W, 60 sec) sessions. Intra-arterial chemotherapy is effective for unresectable metastatic liver tumors from colon cancer. If a patient shows a partial response on the metastatic tumors through the chemotherapy, one must consider other modalities such as PMCT.

[Long-term complete response in a case of unresectable rectal cancer following chemoradiation].

We report a case of unresectable rectal cancer in a 53-year-old male treated with chemoradiation. Radiation therapy was delivered with a total pelvic dose of 45 Gy together with oral administration of 5'-DFUR (1,200 mg/day). The patient received one course of combination chemotherapy consisting of cisplatin, 100 mg/body x 1 day, and 5-FU, 1,000 mg/body x 5 days, followed by radiation therapy. Oral administration of tegafur-uracil (300 mg/day) was continued for five years following the chemoradiation. The patient is now disease-free 75 months after the initial surgery. Chemoradiation can be managed to obtain a complete remission of some locally advanced rectal cancers.

Isolation and characterization of mitochondrial nucleoids from the yeast Pichia jadinii.

Mitochondrial (mt) nucleoids were isolated with a high degree of purity from the yeast Pichia jadinii, in which the mitochondrial DNA (mtDNA) is linear. Field-inversion gel electrophoresis (FIGE) revealed that significant amounts of mtDNA could be isolated intact, as linear molecules of 41 kbp, from the isolated mt-nucleoids. Fifteen different proteins were detected in the mt-nucleoid fraction and, eight of these proteins bound to DNA. The patterns of mt-nucleoid proteins and of the DNA-binding proteins after gel electrophoresis in the presence of SDS were somewhat different from those of such proteins from Saccharomyces cerevisiae. The corresponding proteins isolated from the mt-nucleoids of four other species of yeast in the genera Pichia and Williopsis also differed from one another in terms of electrophoretic mobility in the presence of SDS. In immunoblotting experiments, antibodies that had been raised against the 67-kDa protein of mt-nucleoids from S.cerevisiae and the YMN-1 monoclonal antibody that is specific for a 48-kDa protein in the mt-nucleoids from S. cerevisiae did not recognize any proteins in the mt-nucleoids from Pichia jadinii and four other species of yeast. The results suggest the considerable diversity of the proteins in the mt-nucleoids of yeasts.

Characterization of DNA-binding proteins involved in the assembly of mitochondrial nucleoids in the yeast Saccharomyces cerevisiae.

Mitochondrial nucleoids (mt-nucleoids) isolated from the yeast Saccharomyces cerevisiae were analyzed to identify the protein components that are involved in the compact packaging of mtDNA. The isolated mt-nucleoids were disassembled by the addition of 2 M NaCl and the disassembled mt-nucleoids were reassembled once again into compact structures by dialysis against a buffer that contained NaCl at concentrations below 0.1 M, as monitored by staining of the DNA with 4',6-diamidino-2-phenylindole. DNA-binding proteins with molecular masses of 67 kDa, 52 kDa, 50 kDa, 38 kDa, 30 kDa and 20 kDa were separated from isolated mt-nucleoids by column chromatography on DNA-cellulose after digestion of mt-nucleoids by DNase I in the presence or absence of 2 M NaCl. Purified mtDNA was compactly packaged into nucleoid-like structures upon the addition of fractions that contained DNA-binding proteins and subsequent dialysis to reduce the concentration of NaCl. Five proteins, with molecular masses of 67 kDa, 52 kDa, 50 kDa, 38 kDa and 30 kDa, respectively, had lower affinity for double-stranded DNA than that of the 20-kDa protein. The fraction that contained the five DNA-binding proteins other than the 20-kDa protein was also able to fold mtDNA compactly into nucleoid-like structures. By contrast, the combination of the 20-kDa protein and mtDNA resulted in formation of less tightly packed, string-of-bead structures. These results suggest that at least six different DNA-binding proteins are involved in the organization of the mt-nucleoids.

The natural history of heterotopic ossification in patients who have fibrodysplasia ossificans progressiva. A study of forty-four patients.

Forty-four patients who had fibrodysplasia ossificans progressiva responded by mail to a questionnaire regarding the age at the onset of heterotopic ossification at fifteen commonly involved anatomical sites. The average age of the patients when they responded to the questionnaire was twenty-seven years (range, three to sixty-nine years). The average age at the onset of ossification was five years (range, birth to twenty-five years). The most common sites of early heterotopic ossification were the neck, spine, and shoulder girdle. Thirty-five (80 per cent) of the patients had had some restrictive heterotopic ossification by the age of seven years. By the age of fifteen years, forty-two (more than 95 per cent) of the patients had severely restricted mobility of the upper limbs. In these patients, heterotopic ossification proceeded in a direction that was axial to appendicular, cranial to caudad, and proximal to distal; this pattern appeared typical for fibrodysplasia ossificans progressiva.

Preparation of a monoclonal antibody specific for a 48-kDa protein from mitochondrial nucleoids of the yeast, Saccharomyces cerevisiae.

Monoclonal antibodies (mAbs) were raised against yeast mitochondrial nucleoids (mt-nucleoids). In an analysis by a combination of immunofluorescence microscopy and staining with 4',6-diamidino-2-phenylindole (DAPI), one of them, designated YMN-1, distinctly stained mt-nucleoids, which were visible as dots, in spheroplasts and in isolated mitochondria. However, staining of isolated mt-nucleoids was rather weak. YMN-1 mAb recognized a 48-kDa protein in immunoblots of both mitochondrial and mt-nucleoid proteins. The 48-kDa protein was a minor component of mt-nucleoid proteins and was separated from extract of both mitochondria and mt-nucleoids by immunoaffinity chromatography. The affinity-purified 48-kDa protein reassociated with mt-nucleoids when mixed with isolated mt-nucleoids, as monitored by immunofluorescence microscopy. The results suggest that a large amount of 48-kDa protein is associated with mt-nucleoids in vivo, and that lysis of mitochondria by the treatment with detergent releases a considerable amount of this protein from mt-nucleoids during the isolation of mt-nucleoids.

"Alternative self-diploidization" or "ASD" homothallism in Saccharomyces cerevisiae: isolation of a mutant, nuclear-cytoplasmic interaction and endomitotic diploidization.

A mutant of Saccharomyces cerevisiae representing a novel life cycle, named "alternative self-diploidization" or "ASD" homothallism, was obtained fortuitously. In this life cycle, MAT alpha (or MATa) haplophase and MAT alpha/MAT alpha (or MATa/MATa) diplophase alternate. Germinated cells are haploid and mating. They soon become nonmating and sporogenous as they vegetatively grow. They sooner or later diploidize presumably via endomitosis. The diploid cells haploidize via normal meiosis. A single recessive nuclear mutation, named asd 1-1, is responsible for "ASD" homothallism. In the rho 0 cytoplasm, asd 1-1 cells mate even if at a low efficiency and fail to diploidize. Since pet mutations do not have such effects, we conclude that a certain mitochondrial function other than respiration is required for manifestation of "ASD" homothallism. That is, "ASD" homothallism is the result of some sort of nuclear-cytoplasmic interaction.

Isolation of morphologically intact mitochondrial nucleoids from the yeast, Saccharomyces cerevisiae.

Mitochondrial nucleoids (mt-nucleoids) of the yeast, Saccharomyces cerevisiae, were isolated from spheroplasts of stationary phase cells and their structure and organization were investigated by fluorescence microscopy, electron microscopy, and biochemical techniques. Isolated mt-nucleoids were spherical or ovoid and 0.3-0.6 micron in diameter, and were about the same size and shape as those observed in the cell by the DAPI staining technique. Measurement of DNA content of mt-nucleoids, using a video-intensified microscope system, after DAPI staining revealed that a mt-nucleoid in spheroplasts from stationary phase cells contains, on average, 3.9 mtDNA molecules and an isolated mt-nucleoid contains, on average, 3.1. Negatively stained electron micrographs showed that mt-nucleoids consist of particles 20-50 nm in diameter. SDS-polyacrylamide gel electrophoresis of mt-nucleoids detected 20 species of polypeptides in the molecular weight range from 10 X 10(3) to 70 X 10(3). Acid-urea/SDS two-dimensional electrophoresis of acid extract from mt-nucleoids indicated that a polypeptide of 20 X 10(3) is the only major polypeptide with basic property like histones.

Cytological characterization of NOR in the bivalent of Saccharomyces cerevisiae.

We cytologically characterized the nucleolar organizer region (NOR) on the bivalent in the yeast Saccharomyces cerevisiae. We used staining with 4'-6-diamidino-2-phenylindole (DAPI), chromomycin A3, and silver nitrate and in situ hybridization technique and utilized a video-intensified microscope system with an ultra-high-sensitive video camera. The results showed that of 16 bivalents of S. cerevisiae, the longest was a recognizable nucleolar chromosome which has an annular and synaptonemal complexless NOR in its submedian portion. The NOR was comprised of 2.65 X 10(9) D DNA which corresponded to 118 copies per haploid of rDNA repeating units. This evidence is discussed in terms of the possible participation of the annular NOR in suppressing the meiotic recombination of the rDNA gene clusters.

Meiotic karyotype of the yeast Saccharomyces cerevisiae.

A cytogenetic study of the meiotic chromosomes of the budding yeast Saccharomyces cerevisiae was undertaken by high resolution epifluorescence microscopy. Condensation of chromatin into separate chromosomes takes place during prophase I. At metaphase I, there are 16 separate and distinct bivalents which are roughly classified into three groups by morphological differences and DNA content.

Fluorescence microscopic studies of mitochondrial nucleoids during meiosis and sporulation in the yeast, Saccharomyces cerevisiae.

Configurational changes of mitochondria and mitochondrial nucleoids (mt-nucleoids) during meiosis and sporulation in the yeast, Saccharomyces cerevisiae, were examined using the mitochondrial membrane-binding fluorescent dye, dimethyl aminostyrylmethylpyridiniumiodine (DASPMI) and the DNA-binding fluorescent dye, 4',6-diamidino-2-phenylindole (DAPI). In zygotes just after mating, mt-nucleoids were observed as many small discrete light spots in the cytoplasm. During meiosis in zygotes, mt-nucleoids at first coalesced with each other into a long string and then separated into spherical nucleoids in four spores. These changes paralleled those in mitochondria observed using DASPMI. The use of spheroplasts allowed us to examine the behaviour of mt-nucleoids at higher resolution and to identify several distinct meiotic prophase stages of the cell nucleus during early sporulation. In diploid spheroplasts at the stationary phase, 50-70 of the mt-nucleoids were observed to be separated from each other and each spherical mitochondrion contained only one mt-nucleoid. At the later stage of premeiotic DNA synthesis, a single branched giant mitochondrion was formed as a result of complete mitochondrial fusion. All of the mt-nucleoids were arranged in an array on a giant mitochondrion and coalesced into a string-like network. Through meiosis I and II, strings of mt-nucleoids were observed close to the dividing nuclei. At late meiosis II, a ring of mt-nucleoids enclosing each daughter nucleus was formed. In ascospores, discrete small nucleoids were visible close to each spore nucleus with a 'string-of-beads' appearance. Many mt-nucleoids were excluded from the ascospores and remained in the residual cytoplasm of the ascus.