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Supramolecular drug carriers are a promising approach for delivering anticancer drugs with high blood retention after administration. We previously synthesized folic acid-modified methyl-β-cyclodextrin (FA-MβCD) as an anticancer drug. FA-MβCD has a selective autophagy-mediated antitumor effect on folic acid receptor (FR)-expressing cancer cells. Here, we enhanced the antitumor effect and safety of FA-MβCD by preparing a supramolecular nanoparticle formulation of FA-MβCD via host-guest interactions using an adamantane conjugate with human serum albumin (Ad-HSA). The Ad-HSA/FA-MβCD supramolecular complex prolonged the blood retention of FA-MβCD and improved its antitumor effect and safety after intravenous administration in tumor-bearing mice xenografted with FR-expressing cancer cells. These results suggest that the supramolecular technique using Ad-HSA is a promising approach for the delivery of CD-based anticancer drugs.

Hepatitis B (HB) vaccines (Heptavax-II and Bimmugen) designed based on HBV genotypes A and C are mainly used for vaccination against HB in Japan. To determine whether there are differences in the genetic background associated with vaccine responsiveness, genome-wide association studies were performed on 555 Heptavax-II and 1193 Bimmugen recipients. Further HLA imputation and detailed analysis of the association with HLA genes showed that two haplotypes, DRB1*13:02-DQB1*06:04 and DRB1*04:05-DQB1*04:01, were significantly associated in comparison with high-responders (HBsAb > 100 mIU/mL) for the two HB vaccines. In particular, HLA-DRB1*13:02-DQB1*06:04 haplotype is of great interest in the sense that it could only be detected by direct analysis of the high-responders in vaccination with Heptavax-II or Bimmugen. Compared with healthy controls, DRB1*13:02-DQB1*06:04 was significantly less frequent in high-responders when vaccinated with Heptavax-II, indicating that high antibody titers were less likely to be obtained with Heptavax-II. As Bimmugen and Heptavax-II tended to have high and low vaccine responses to DRB1*13:02, 15 residues were found in the Heptavax-II-derived antigenic peptide predicted to have the most unstable HLA-peptide binding. Further functional analysis of selected hepatitis B patients with HLA haplotypes identified in this study is expected to lead to an understanding of the mechanisms underlying liver disease.

We report a patient with autism and cleft lip and palate carrying a de novo heterozygous AUTS2 mutation, c.1464_1467del ACTC (p.Tyr488*). Although the causal relationship between cleft lip and palate and this mutation is unclear, this case report may expand the clinical phenotype of AUTS2 syndrome.

Objective Since healthcare providers face an increased risk of hepatitis B virus (HBV) infection because of their work, vaccination plays a critical role in preventing HBV transmission. However, the duration for which acquired HBV surface antibodies (anti-HBs) persist remains unknown. To evaluate the primary immunologic response to HBV vaccination and its persistence in healthy Japanese adolescents. Methods In total, 690 young adults underwent HBV vaccination with a three-dose schedule. The primary response was determined by the anti-HBs titers at 1-2 months after the final dosage. Subjects with anti-HBs titers of 100 mIU/mL were classified as “non-responders,” “low-responders,” and “sufficient responders,” respectively. Anti-HB titers were re-measured at 1 or 2 years after vaccination. Results First, 95.8% and 72.8% of the subjects had anti-HBs titers of >10 and >100 mIU/mL, respectively, as a primary response. The anti-HBs titers measured at 1 and 2 years after vaccination were significantly correlated with those of the primary response (1 year: r=0.893, p100 mIU/mL at the primary response maintained an anti-HBs titer of >10 mIU/mL [1 year after vaccination, 208/209 (99.5%); 2 years after vaccination, 72/81 (90.1%)]. However, in subjects with a primary response of 10-100 mIU/mL the anti-HBs titer frequently declined; 17/38 (44.7%) and 9/10 (90.0%) subjects had a titer of

Approximately 5-10% of individuals who are vaccinated with a hepatitis B (HB) vaccine designed based on the hepatitis B virus (HBV) genotype C fail to acquire protective levels of antibodies. Here, host genetic factors behind low immune response to this HB vaccine were investigated by a genome-wide association study (GWAS) and Human Leukocyte Antigen (HLA) association tests. The GWAS and HLA association tests were carried out using a total of 1,193 Japanese individuals including 107 low responders, 351 intermediate responders, and 735 high responders. Classical HLA class II alleles were statistically imputed using the genome-wide SNP typing data. The GWAS identified independent associations of HLA-DRB1-DQB1, HLA-DPB1 and BTNL2 genes with immune response to a HB vaccine designed based on the HBV genotype C. Five HLA-DRB1-DQB1 haplotypes and two DPB1 alleles showed significant associations with response to the HB vaccine in a comparison of three groups of 1,193 HB vaccinated individuals. When frequencies of DRB1-DQB1 haplotypes and DPB1 alleles were compared between low immune responders and HBV patients, significant associations were identified for three DRB1-DQB1 haplotypes, and no association was identified for any of the DPB1 alleles. In contrast, no association was identified for DRB1-DQB1 haplotypes and DPB1 alleles in a comparison between high immune responders and healthy individuals. Conclusion: The findings in this study clearly show the importance of HLA-DR-DQ (i.e., recognition of a vaccine related HB surface antigen (HBsAg) by specific DR-DQ haplotypes) and BTNL2 molecules (i.e., high immune response to HB vaccine) for response to a HB vaccine designed based on the HBV genotype C. (Hepatology 2018).

Although an inhibition of action potential conduction in nerve fibers possibly contributes to at least a part of antinociception produced by analgesics and the adjuvants, it has not been fully examined yet how the conduction inhibition differs in extent among their drugs. We investigated the effects of various antidepressants used as analgesic adjuvants on compound action potentials (CAPs) recorded from the frog sciatic nerve by using the air-gap method. The results were compared with those of the other adjuvants that were reported previously. Antidepressants, duloxetine (serotonin and noradrenaline reuptake inhibitor, SNRI), fluoxetine (selective serotonin reuptake inhibitor, SSRI), amitriptyline (tricyclic tertiary amine), desipramine (tricyclic secondary amine) and maprotiline (tetracyclic secondary amine), reduced the peak amplitude of the CAP with half-maximal inhibitory concentration (IC50) values of 0.23, 1.5, 0.26, 1.6 and 0.95mM, respectively. Trazodone (non-SNRI, -SSRI, -tricyclic and -tetracyclic antidepressant) at 1.0mM reduced CAP amplitude by about 50%. The duloxetine and amitriptyline values were comparable to those of lamotrigine and carbamazepine (antiepileptics), dexmedetomidine (α2-adrenoceptor agonist) and ropivacaine, levobupivacaine and pramoxine (local anesthetics). The fluoxetine, desipramine, maprotiline and trazodone values were similar to those of oxymetazoline (α2-adrenoceptor agonist) and lidocaine, cocaine, procaine and prilocaine (local anesthetics). The antidepressants' IC50 values were much larger than that of tetracaine (local anesthetic). In conclusion, the six antidepressants inhibited CAPs with efficacies comparable to some antiepileptics, α2-adrenoceptor agonists and local anesthetics. It was suggested that antidepressants inhibit nerve conduction with efficacies comparable to those of the other adjuvants.

Phosphatase and tensin homolog (PTEN) plays an important role in tumor suppression. A germline mutation in the PTEN gene induces not only PTEN hamartoma tumor syndrome, including Cowden syndrome, but also macrocephaly/autism syndrome. Here, we describe a boy with macrocephaly/autism syndrome harboring a novel missense heterozygous PTEN mutation, c.959T>C (p.Leu320Ser). Interestingly, a previously reported nonsense mutation resulting in p.Leu320X was found in Cowden syndrome patients. Our case may be suggestive of a genotype-phenotype correlation.

Phosphatase and tensin homolog (PTEN) plays an important role in tumor suppression. A germline mutation in the PTEN gene induces not only PTEN hamartoma tumor syndrome, including Cowden syndrome, but also macrocephaly/autism syndrome. Here, we describe a boy with macrocephaly/autism syndrome harboring a novel missense heterozygous PTEN mutation, c.959T>C (p.Leu320Ser). Interestingly, a previously reported nonsense mutation resulting in p.Leu320X was found in Cowden syndrome patients. Our case may be suggestive of a genotype-phenotype correlation.

The pharmacokinetics among children has been altered dynamically. The difference between children and adults is caused by immaturity in things such as metabolic enzymes and transport proteins. The periods when these alterations happen vary from a few days to some years after birth. We hypothesized that the effect of gene polymorphisms associated with the dose of medicine could be influenced by age. In this study, we analyzed 51 patients with childhood acute lymphoblastic leukemia (ALL) retrospectively. We examined the associations between the polymorphism in NUDT15 and clinical data, especially the dose of 6-mercaptopurine (6-MP). Ten of the patients were heterozygous for the variant allele in NUDT15. In patients under 7 years old with NUDT15 variant allele, the average administered dose of 6-MP was lower than that for the patients homozygous for the wild-type allele (P=0.04). Genotyping of NUDT15 could be a beneficial to estimate the tolerated dose of 6-MP for patients with childhood ALL, especially at a preschool age in Japan. Furthermore, the analysis with stratification by age might be useful in pharmacogenomics among children.

Background Food allergy is a growing health problem worldwide because of its increasing prevalence, life-threatening potential, and shortage of effective preventive treatments. In an outbreak of wheat allergy in Japan, thousands of patients had allergic reactions to wheat after using soap containing hydrolyzed wheat protein (HWP). Objectives The aim of the present study was to investigate genetic variation that can contribute to susceptibility to HWP allergy. Methods We conducted a genome-wide association study of HWP allergy in 452 cases and 2700 control subjects using 6.6 million genotyped or imputed single nucleotide polymorphisms. Replication was assessed by genotyping single nucleotide polymorphisms in independent samples comprising 45 patients with HWP allergy and 326 control subjects. Results Through the genome-wide association study, we identified significant associations with the class II HLA region on 6p21 (P = 2.16 × 10−24 for rs9271588 and P = 2.96 × 10−24 for HLA-DQα1 amino acid position 34) and with the RBFOX1 locus at 16p13 (rs74575857, P = 8.4 × 10−9). The associations were also confirmed in the replication data set. Both amino acid polymorphisms (HLA-DQβ1 amino acid positions 13 and 26) located in the P4 binding pockets on the HLA-DQ molecule achieved the genome-wide significance level (P Conclusions Our data provide the first demonstration of genetic risk for HWP allergy and show that this genetic risk is mainly represented by multiple combinations of HLA variants.

Aim To assess the feasibility of transferring to the University of Tsukuba Hospital for proton beam therapy (PBT) during intensive chemotherapy in children with Ewing sarcoma family of tumors (ESFT) who had been diagnosed and started their first-line treatment at prefectural or regional centers for pediatric oncology. Background The treatment of ESFT relies on a multidisciplinary approach using intensive neoadjuvant and adjuvant chemotherapies with surgery and radiotherapy. Multi-agent chemotherapy comprising vincristine, doxorubicin, cyclophosphamide, ifosfamide, and etoposide (VDC-IE) is widely used for ESFT, and the interval between each course is very important for maintaining the intensity and effect of chemotherapy. Materials and methods Clinical information of patients who received PBT and VDC-IE between April 2009 and May 2016 was collected retrospectively. The intervals between each course of VDC-IE and adverse events were assessed. Results Fifteen patients were evaluated. No delays in the intervals of chemotherapy due to transfer were observed. There were no adverse events caused during/just after transfer and no increases in adverse events. The estimated 4-year overall and event-free survival rates were 94.6% and 84.8%, respectively. Discussion Although the results of efficacy are preliminary, survival rates were comparable with past studies. More experience and follow-up are required to further assess the efficacy of PBT for patients with ESFT. Conclusion Multidisciplinary therapy for children with ESFT involving transfer to our hospital for PBT during VDC-IE was feasible without treatment delay or an increase in adverse events.

Imatinib mesylate has dramatically improved the outcome of children with Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph* ALL) and is now included as first-line therapy. Uncommon adverse effects of this drug for pediatric use, however, are largely unknown. We report the first case of a 9-year-old child who developed severe acute hepatitis with grade 4 transaminases and bilirubin elevation during imatinib treatment for Ph* ALL. Liver biopsy showed extensive lobular and pericentral necrosis of hepatocytes. Liver function recovered after discontinuing imatinib with a 4-week prednisolone. Extensive hepatic necrosis should be considered not only in adults but also in children under imatinib administration.

Background Management of the adverse effects of chemotherapy is essential to improve outcome of children with leukemia. Some genetic polymorphisms can predict treatment-related toxicity, and be used individually in dose modification of 6-mercaptopurine (6-MP) and methotrexate (MTX) in maintenance therapy for childhood acute lymphoblastic leukemia (ALL). We investigated associations between clinical course and candidate gene polymorphisms less evaluated in Japanese patients. Methods Fifty-three children who received maintenance chemotherapy were enrolled in this study. The scheduled dose of oral 6-MP was 40 mg/m2 daily and that of oral MTX was 25 mg/m2 weekly. The doses were adjusted according to white blood cell count (target range, 2.5–3.5 × 109/L) and aspartate aminotransferase and alanine aminotransferase level ( C than in the patients with wild homozygous genotype. The other analyzed polymorphisms were not associated with toxicity, 6-MP, or MTX dose. Conclusions Polymorphism of SLCO1B1 c.521 T > C could be a strong predictor of 6-MP dose reduction in maintenance chemotherapy in childhood ALL.

We report a 10-year-old girl with Bardet–Biedl syndrome caused by a novel mutation in the Bardet–Biedl syndrome 10 (BBS10) gene. She had multiple malformations, including a dysmorphic face, postaxial polydactyly, polycystic kidney and amblyopia. She presented with typical BBS features, including intellectual disability with emotional outbursts and mild obesity. Whole-exome sequencing identified compound heterozygous mutations with NM_024685.3:c.1677C>A [p.(Tyr559*)] and c.1974T>G [p.(Tyr658*)]. To our knowledge, the latter mutation has never been reported previously.

Wiedemann-Steiner syndrome (WDSTS) is an autosomal dominant disorder characterized by hypertrichosis, intellectual disability, and dysmorphic facial appearances (down-slanted vertically narrow palpebral fissures, wide nasal bridge, broad nasal tip, and thick eyebrows). In 2012, Jones and co-workers identified heterozygous mutations in KMT2A (lysine methyltransferase 2A) as the molecular cause of WDSTS. Although the phenotype of this syndrome continues to expand, the associated features are not fully understood. Here, we report WDSTS in a 12-year-old Japanese boy with a novel nonsense mutation in KMT2A. He had right preaxial polydactyly, which has not been previously reported in WDSTS. We could not identify a causal relationship between the KMT2A mutation and preaxial polydactyly, and cannot exclude the preaxial polydactyly is a simple coincidence. We summarized the clinical features of WDSTS associated with KMT2A mutation and discussed the cardinal symptoms in detail.

We report a 10-year-old girl with Bardet–Biedl syndrome caused by a novel mutation in the Bardet–Biedl syndrome 10 (BBS10) gene. She had multiple malformations, including a dysmorphic face, postaxial polydactyly, polycystic kidney and amblyopia. She presented with typical BBS features, including intellectual disability with emotional outbursts and mild obesity. Whole-exome sequencing identified compound heterozygous mutations with NM_024685.3:c.1677C>A [p.(Tyr559*)] and c.1974T>G [p.(Tyr658*)]. To our knowledge, the latter mutation has never been reported previously.

Backgrounds. Outcome of childhood malignancy has been improved mostly due to the advances in diagnostic techniques and treatment strategies. While methotrexate (MTX) related polymorphisms have been under investigation in childhood malignancies, many controversial results have been offered. Objectives. To evaluate associations of polymorphisms related MTX metabolisms and clinical course in childhood lymphoid malignancies. Method. Eighty-two acute lymphoblastic leukemia and 21 non-Hodgkin’s lymphoma children were enrolled in this study. Four single nucleotide polymorphisms in 2 genes (MTHFR (rs1801133/c.677C>T/p.Ala222Val and rs1801131/c.1298A>C/p.Glu429Ala) and SLCO1B1 (rs4149056/c.521T>C/p.V174A and rs11045879/c.1865+4846T>C)) were genotyped by Taqman PCR method or direct sequencing. Clinical courses were reviewed retrospectively. Results. No patient who had the AC/CC genotype of rs1801131 (MTHFR) had relapsed or died, in which distribution was statistically different among the AA genotype of rs1801131 (P=0.004). Polymorphisms of SLCO1B1 (rs11045879 and rs4149056) were not correlated with MTX concentrations, adverse events, or disease outcome. Conclusions. Polymorphisms of MTHFR (rs1801131) could be the plausive candidate for prognostic predictor in childhood lymphoid malignancies.

Background & aims Genetic factors in class II human leukocyte antigen ( HLA ) have been reported to be associated with inter-individual variation in hepatitis B virus (HBV) vaccine response. However, the mechanism underlying the associations remains elusive. In particular, the broad linkage disequilibrium in HLA region complicates the localization of the independent effects of genetic variants. Thus, the present study aimed to identify the most probable causal variations in class II HLA loci involved in the immune response to HBV vaccine. Methods We performed a case-control study to assess whether HLA-DRB1 , - DQB1 , and -DPB1 4-digit alleles were associated with the response to primary HBV vaccination in 574 healthy Japanese students. To identify causative variants, we next assessed independently associated amino acid variants in these loci using conditional logistic regression analysis. Furthermore, to clarify the functional effects of these variants on HLA proteins, we performed computational structural studies. Results HLA-DRB1 ∗ 01:01 , HLA-DRB1 ∗ 08:03 , HLA-DQB1 ∗ 05:01 , and HLA-DPB1 ∗ 04:02 were significantly associated with sufficient response, whereas HLA-DPB1 ∗ 05:01 was associated with poor response. We then identified amino acids independently associated with sufficient response, namely, leucine at position 26 of HLA-DRβ1 and glycine-glycine-proline-methionine at positions 84–87 of HLA-DPβ1. These amino acids were located in antigen-binding pocket 4 of HLA-DR and pocket 1 of HLA-DP, respectively, which are important structures for selective binding of antigenic peptides. In addition, the detected variations in HLA-DP protein were responsible for the differences in the electrostatic potentials of the pocket, which can explain in part the sufficient/poor vaccine responses. Conclusion HLA-DRβ1 position 26 and HLA-DPβ1 positions 84–87 are independently associated with anti-HBs production against HBV vaccine. Our results suggest that HBsAg presentation through these HLA pocket structures plays an important role in the inter-individual variability of HBV vaccination.

Although there is ample evidence supporting the effectiveness of physical activity in the prevention and treatment of osteoporosis, there are no previous studies to examine the effect of office-based brief high-impact exercise (HIE) on bone mineral density (BMD) in healthy premenopausal women. This study evaluated the effects of office-based HIE on BMD in healthy premenopausal Japanese women. Ninety-one healthy premenopausal women were randomized to receive stretching exercise (SE) or HIE (stretching, along with up to 5 × 10 vertical and versatile jumps) for 12 months. The BMD of the lumbar spine and proximal femur was measured using dual-energy X-ray absorptiometry. Several cardiovascular risk factors and leg strength also were assessed. An accelerometer-based recorder was used to measure daily impact loading in four 1-week samples. The progression of the HIE program was ensured by the accelerometer. Thirty-three women (71.7%) in the SE group and 34 (75.6%) in the HIE group completed the study. There was a significant difference in the change in the femoral neck BMD between the groups in favor of the HIE group [0.6% (95% CI: -0.4, 1.7) vs. -1.0% (95% CI: -2.2, 0.2)]. Adiponectin, LDL, HDL, and the leg strength of participants in both the groups improved during the intervention. These finding suggested that office-based brief HIE can be recommended for premenopausal women for preventing bone mineral loss.

Management of the adverse effects of chemotherapy is essential to improve outcome of children with leukemia. Some genetic polymorphisms can predict treatment-related toxicity, and be used individually in dose modification of 6-mercaptopurine (6-MP) and methotrexate (MTX) in maintenance therapy for childhood acute lymphoblastic leukemia (ALL). We investigated associations between clinical course and candidate gene polymorphisms less evaluated in Japanese patients.Fifty-three children who received maintenance chemotherapy were enrolled in this study. The scheduled dose of oral 6-MP was 40 mg/m(2) daily and that of oral MTX was 25 mg/m(2) weekly. The doses were adjusted according to white blood cell count (target range, 2.5-3.5 × 10(9) /L) and aspartate aminotransferase and alanine aminotransferase level (750 IU/L). Eight polymorphisms in six candidate genes, TPMT, ITPA, MRP4, MTHFR, RFC1, and SLCO1B1, were genotyped using the Taqman PCR method. Clinical course was reviewed retrospectively from medical records.The average dose of 6-MP was lower in the patients with at least one variant allele at SLCO1B1 c.521 TC than in the patients with wild homozygous genotype. The other analyzed polymorphisms were not associated with toxicity, 6-MP, or MTX dose.Polymorphism of SLCO1B1 c.521 TC could be a strong predictor of 6-MP dose reduction in maintenance chemotherapy in childhood ALL.

Rhabdomyosarcoma (RMS) is one of the most common soft tissue sarcomas among children. Patients who developed genitourinary/pelvic rhabdomyosarcoma (GU/P-RMS) have a higher complication ratio and relatively poorer event free survival, with local therapy being very important. While proton beam therapy (PBT) is expected to reduce co-morbidity, especially for children, this lacks firm evidence and analysis. We analyzed GU/P-RMS children who had undergone multimodal therapy combined with PBT at a single institution.We retrospectively reviewed charts of children with GU/P-RMS treated from January 2007 to May 2013 at the University of Tsukuba Hospital who had undergone multimodal therapy with PBT.There were 5 children and their median age at diagnosis was 2.8 years (0.6-4.4 years). Primary sites were the bladder (2) and the prostate (3). All received neo-adjuvant chemotherapy and 3 underwent chemotherapy during PBT (Group Cx). All patients of Group Cx developed leukocytopenia (WBC1000/μL). The median dose of PBT was 47.7 GyE (41.4-50.4 GyE). All patients survived by their last hospital visit (median, 36 months).We analyzed multimodal treatment combined with PBT applied for GU/P-RMS. PBT was well tolerated and could be a plausible choice instead of photon therapy for this population.

The localization of Ca2+ in cells of the periblem and dermatogen in the root meristem and the columella and peripheral cells of the root cap of maize was examined by the precipitation method of potassium pyroantimonate and EGTA-treatment. In periblem and dermatogen cells, Ca2+ was found to be localized in the nucleoplasm and granular zone of the nucleolus. Ca2+ was also found in most cell organelles: in the matrix in mitochondria, on the thylakoid membrane in proplastids, in the vacuoles and on the plasma membranes. Ca2+ was also distributed throughout the cytoplasmic ground matrix. Much Ca2+ was present in the cell wall soon after its formation during the cell division. Ca2+ was also conspicuous in the vesicles of Golgi in the dermatogen cells. In columella and peripheral cells, there was less Ca2+ in the organelles and cytoplasmic ground matrix, but Ca2+ was present in Golgi vesicles in the peripheral cells. Electron microscopic and X-ray microanalysis showed that Ca2+ was also present in the mucilaginous layer, the outermost cell wall of the peripheral cells.

The distribution of Ca2+ in dividing cells of the maize root tip was examined by potassium pyroantimonate precipitation and EGTA treatment methods. Ca2+ was found in most of the cell organelles, such as the matrix of the mitochondria, the thylakoid membrane of the proplastid and the Golgi vesicles, and on the plasma membrane. Ca2+ was also distributed throughout the cytoplasmic ground matrix and attractoplasm, inside the vacuoles, in the granular zone of the nucleolus in the interphasic nucleus and in the regenerated nucleolus in the telophasic nucleus. The amounts of Ca2+ distributed in the cytoplasmic ground matrix, the vacuole and the nucleolus varied during nuclear division. From the results of the present experiment, the following considerations on the role of Ca2+ and the regulation site of Ca2+ in dividing plant cells were drawn: 1) Ca2+ may play a role in the construction of the granular form of the ribosome. 2) Ca2+ may be an essential ion in the regeneration of nucleolus. 3) Vacuoles may act as the regulatory site of the Ca2+ concentration in the cytoplasm and attractoplasm in plant cells. Spindle microtubules and phragmo-microtubules are probably surrounded by other ions, such as Mg2+.

Centriolar plaque and spindle microtubules in young asci of an ascomycete,Sordaria humana were studied by electron microscopy. Centriolar plaque is electron opaque and has an amorphous structure. Two dispositions of centriolar plaque were observed, one entirely contiguous to the nuclear envelope in a meiotic division and the other partially joined to the envelope in a mitotic division following meiosis. The spindle was formed inside the nuclear envelope and spindle microtubules terminated at the polar protrusion of the nuclear envelope. Some spindle microtubules seem to connect directly with the centriolar plaque passing through perforations of the nuclear envelope.

Light and electron microscope observations on resting and dividing nuclei in mycelia, conidiophores and ungerminated and germinated conidia of Erysiphe pisi , after fixation and staining with aceto-orcein and lead citrate, are described. The observations revealed that mitosis is similar to that in other fungi. During early stages of division the nucleolus was clearly visible. The chromosome number was n = 6 but there was little difference in morphology of the chromosomes. The nucleus and nucleolus were clearly visible under the electron microscope.

The origin and the formation of the ascospore delimiting membrane in Sordaria humana were studied by electron microscopy. The complete delimiting membrane consists of a double unit membrane system. It originates from tubular elements of 70–100 nm diameter which are formed with a thin membrane 5–6 nm thick and filled with electron translucent materials. The process of development of the delimiting membrane is as follows: Tubular elements first aggregate, and then become arranged in a plane. Soon afterwards, they fuse laterally and become transformed into a delimiting membrane. Spore delimitation starts from the distal end of the centriolar plaque and develops towards the opposite side of the nucleus, preceding the formation of the membrane system.

Studies were made on the fine structure and chemical components of the synaptonemal complex (SC) of pollen mother cells of Tradescantia reflexa. The SC of T. reflexa consists of three partite structures: a central element consisting of two tubules of about 20 nm width, which are twisted together, a lateral element appearing as an electron-opaque line of 25-35 nm width, and transverse filaments of about 10 nm width which connect the central and lateral elements.The central element contains basic protein and a small amount of DNA and RNA. The lateral element contains basic protein and small amounts of DNA and RNA. The transverse filaments contain non-basic protein.

Electron microscopic studies on the substructure of chromatin were carried out using chromosomes in the first meiotic metaphase from pollen mother cells of Tradescantia reflexa. The chromosomes were composed of 300A chromatin threads formed by coiling of 100A chromatin thread. The 100A chromatin threads contained globular structures of about 70A diameter. The location of DNA, protein and RNA were studied by treatment of the chromosomes with DNAase, 0.25N HCl containing 2M NaCl and RNAase.

Cytochemical studies were made on the kinetochore of chromosomes in the first meiotic metaphase and anaphase of pollen mother cells of Tradescantia reflexa. The kinetochore of T. reflexa is ball- or cup-shaped and about 700 nm in diameter. It seems to contain DNA, RNA and basic proteins judging from the effect of uranyle-EDTA treatment, RNase digestion and PTA-staining. The location of these chemical components were also examined.

The fine structure and ATPase activity of the mitotic spindle in neoblasts of planaria were examined. In neoblasts, the cells have a large nucleus and nucleolus. Mitochondria are aggregated around the nucleus with chromatoid bodies adjacent. The cytoplasm contains little endoplasmic reticulum (ER) and few Golgi bodies but many free ribosomes, forming polysomes, can be seen throughout the cytoplasmic and spindle ground areas. In addition, centriolar bodies, atypical centrioles, can also be recognized in the cytoplasm. Cells in the G2 stage contain a pair of electron-dense bodies, both consisting of fibrogranules but differing from each other in fine structure and, in the mitotic stage, only one fibrogranular body can be recognized at each pole. ATPase activity was detected in the centriolar bodies in the G2 and mitotic stages and in the ground area of the cytoplasm and spindle apparatus filled by free ribosomes. The activity associated with the microtubules differed with the developmental stage.

The development of achrosomes in spermiogenesis of Blattella germanica was studied by electron microscopy. Achrosomes consist of an achrosomal vesicle originating from Golgi vesicles and an axial rod composed of fine fibrils. The achrosomal vesicle, formed at the mature face of the Golgi body, migrates to the anterior of the nucleus, where it later becomes the front structures of sperm head. After attachment to the nucleus, the achrosomal vesicle changes from a round to a tapering shape, passing through a coneshape phase. During these changes, the axial rod develops in the hollow formed by indentations of adjacent parts of the achrosomal vesicle and the nucleus. The cisternae of the Golgi body concerned with formation of the achrosomal vesicle, are made by pinching off small vesicles from both the ER and the nuclear envelope.

The course of formation of the stack of annulate lamellae was studied in spermatogonial cells of the grasshopper, Atractomorpha bedeli. A tubular body with a very compact and regular pattern occurs often in the cytoplasm (Fig. 1). Some tubules in the tubular body protrude into the cytoplasm, becoming wider in diameter (Figs. 2 and 3). Protruding tubules may fuse with each other forming a lamella (Figs. 2 and 3). As a result, many parallel cisternae with an annuli or a periodic pattern (Fig. 4) develop in the form of a stack of annulate lamellae. With the further growth of the lamellar stack the compact and regular form of the tubular body is lost (Fig. 6). The tubular body gradually becomes smaller (Figs. 7 and 8), and finally disappears when the lamellar stack has developed fully (Fig. 9).

The behavior of mitochondria was studied in pollen mother cells of Tradiscantia reflexa fixed with OsO4 or KMnO4 under the electron microscope. Mitochondria were found scattered throughout the cytoplasm in cells in prophase I. But with the break down of the nuclear membrane (prometaphase I), mitochondria were found intruded into the nuclear or spindle area. Besides mitochondria, dictyosomes, lipid granules and even plastids were seen in the spindle. Mitochondria in the spindle area or phragmoplast gradually increased in number with the progress of meiotic stages from metaphase I to early telophase I. In mid-telophase I, most of the mitochondria are aggregated about the equator of the phragmoplast. Vesicles were found among the aggregated mitochondria arranged in a series forming the beginning of the cell plate (late telophase I). By the cell plate formed, the mitochondria are distributed evenly between the daughter cells. The relation between the increase in elements of the endoplasmic reticulum at the boundary of the spindle followed by the break down of the nuclear membrane and the intrusion of mitochondria into the spindle is discussed.

The course of formation of the stack of cisternal ER and the tubular ER were studied in successive stages of meiotic divisions in Atractomorpha bedeli.A very few stacks of cisternal ER or tubular ER are seen in the cytoplasm of gonial cells (Fig. 1). In cells in diplotene and diakinesis stages (Fig. 15), however, the cytoplasm is filled with agranular tubular ER. In some cells in early meiotic prophase (leptotene stage), agglomeration of vesicles is seen in the vicinity of Golgi bodies or between Golgi bodies (Figs. 4 and 5). In this region, near to the Golgi body or between Golgi bodies, a developing stack of granular ER cisternae is seen in cells in the pachytene stage (Figs. 7-13). The elements in the ER stack are continuous with the Golgi vesicles. The Golgi vesicles furthest outside the Golgi cisternae are oblong and semi-granular, suggesting that they are half-formed ER cisteranae. In these stages, tubular ER are also seen which are limited to the vicinity of the Golgi body, and continuation of tubular ER with the Golgi vesicles is often observed. These facts suggest that the stack of cisteranl ER and tubular ER are formed by fusion of Golgi vesicles. The stack of cisternal ER afterwards moves in the cytoplasm to a place different from the Golgi region and becomes attached to the nuclear envelope (cf. diakinesis stage, Fig. 14).

The fine structure of centrioles was studied in spermatocytes at prophase of vo species grasshoppers, Acrida turrita and Atractomorplia bedeli. It was found iat the centrioles consist of two equal parts: a proximal half (near the nucleus) is a thickness of 50mμ in width and an electron opaque wall, while a distal half is a thinner (25mμ in width) and less electron opaque wall. The distal half of Le centrioles is surrounded by a granular body which appears to originate from tercellular substance.

The development of the spindle body was studied in the spore mother cells of Equisetum arvense under the electron microscope. The spindle body consists of two elements, spindle fibers and a spindle matrix filling the spaces between these spindle fibers. These spindle fibers originate from the cytoplasm as follows. Microtubules of 25mμ in diameter become visible among the cytoplasmic organelles (such as mitochondria and plastids) in early diakinesis (Fig. 2). These microtubules move to the cytoplasmic region around the nucleus (Figs. 3-6) during diakinesis, while other organelles such as mitochondria and plastids move to the periphery of the cell (forming the clear zone). As soon as the continuous nuclear envelope breaks into fragments (prometaphase-I), these microtubules intrude into the nuclear area from the clear zone together with ER, small vesicles (30-100mμ in diameter) and cytoplasmic matrix (Figs. 7 and 8). They may connect with the kinetochore of each chromosome (chromosomal fibers, Fig. 9). Some microtubules not connected with kinetochore may form continuous fibers running from pole to pole (Figs. 9 and 10).