Electron Microscopic Studies of the Differentiation of Fat Cells in Human Fetal Skin**From the Department of Dermatology, Yamaguchi University, School of Medicine, Ube, Japan.

ABSTRACT1)Electron microscopic studies of primitive fat organs showing various degrees of fat storage from human fetuses 20 to 26 weeks of age were undertaken, with special reference to the origin of fat cells and the lipid formations in them. Based on this study, three types of cells, namely an undifferentiated-type cell, a young-type fat cell and a mature-type fat cell were discerned ultrastructurally, but they are considered to be identical concerning the morphology of mitochondria, smooth endoplasmic reticula and glycogen granules.2)Concerning the origin of white fat cells, it was revealed that young-type and mature-type fat cells are not derived from reticuloendothelial cells or fibroblasts but from a certain definite type of mesenchymal cell which we have referred to as the undifferentiated-type cell.3)The finding that cytoplasmic microvesicles are more prominent in young-type fat cells than in mature-type ones might be interpreted that the young-type cells may be actively releasing lipids or free fatty acids according to Williamson's concept of these organelles. But, contrary to Williamson's postulation, the young-type fat cells seem to be accumulating lipids or free fatty acids

N-methyl-D-aspartate receptors play important roles in acquisition and expression of the eyeblink conditioned response in glutamate receptor subunit delta2 mutant mice.

Classical eyeblink conditioning has been known to depend critically on the cerebellum. Apparently consistent with this, glutamate receptor subunit delta2 null mutant mice, which have serious morphological and functional deficiencies in the cerebellar cortex, are severely impaired in delay paradigm. However, these mutant mice successfully learn in trace paradigm, even in \u270-trace paradigm,\u27 in which the unconditioned stimulus starts just after the conditioned stimulus terminates. Our previous studies revealed that the hippocampus and the muscarinic acetylcholine receptors play crucial roles in 0-trace paradigm in glutamate receptor subunit delta2 null mutant mice unlike in wild-type mice, suggesting a large contribution of the forebrain to 0-trace conditioning in this type of mutant mice. In the present study, we investigated the role of N-methyl-D-aspartate receptors in 0-trace eyeblink conditioning in glutamate receptor subunit delta2 null mutant mice. Mice were injected intraperitoneally with the noncompetitive N-methyl-d-aspartate receptor antagonist (+)MK-801 (0.1mg/kg) or saline, and conditioned with 350-ms tone conditioned stimulus followed by 100-ms periorbital shock unconditioned stimulus. Glutamate receptor subunit delta2 null mutant mice that received (+)MK-801 injection exhibited a severe impairment in acquisition of the conditioned response, compared with the saline-injected glutamate receptor subunit delta2 null mutant mice. In contrast, wild-type mice were not impaired in acquisition of 0-trace conditioned response by (+)MK-801 injection. After the injection solution was changed from (+)MK-801 to saline, glutamate receptor subunit delta2 null mutant mice showed a rapid and partial recovery of performance of the conditioned response. On the other hand, when the injection solution was changed from saline to (+)MK-801, glutamate receptor subunit delta2 null mutant mice showed a marked impairment in expression of the pre-acquired conditioned response, whereas impairment of the expression was small in wild-type mice. Injection of (+)MK-801 had no significant effects on spontaneous eyeblink frequency or startle eyeblink frequency to the tone conditioned stimulus in either glutamate receptor subunit delta2 null mutant mice or wild-type mice. These results suggest that N-methyl-D-aspartate receptors play critical roles both in acquisition and expression of the conditioned response in 0-trace eyeblink conditioning in glutamate receptor subunit delta2 null mutant mice

Pembelajaran Berbasis Projek dengan Pendekatan Jelajah Alam Sekitar sebagai Model Perkuliahan Fisiologi Hewan

Penelitian ini dilakukan dengan tujuan untuk menguji efektivitas penerapan model pembelajaran berbasis projek (Project Based Learning/PBL) dengan pendekatan Jelajah Alam Sekitar (JAS) pada perkuliahan Fisiologi Hewan bagi mahasiswa Program Studi Pendidikan Biologi. Penelitian ini merupakan quasi eksperimen dengan desain one shot case study. Populasi dalam penelitian ini adalah mahasiswa Program Studi Pendidikan Biologi dan sebagai sampel adalah mahasiswa semester 4 Prodi Pendidikan Biologi rombel 1,2,dan 3 tahun ajaran 2010 / 2011 yang mengambil mata kuliah Fisiologi Hewan. Pengambilan sampel dengan teknik purposive random sampling. Sebagai variabel bebas adalah penerapan model PBL dengan pendekatan JAS, sedangkan sebagai variabel terikat adalah efektivitas model pembelajaran yang diterapkan dilihat dari hasil belajar, aktivitas siswa dan keterlaksanaan kegiatan yang diprogramkan. Sumber data penelitian adalah mahasiswa. Data yang diambil adalah nilai ujian tulis, nilai laporan, nilai presentasi , aktivitas mahasiswa serta tanggapan keterlaksanaan PBL dengan pendekatan JAS. Hasil penelitian menunjukkan bahwa penerapan model PBL dengan pendekatan JAS telah dapat mencapai indikator-indikator yang ditetapkan yaitu mahasiswa yang memperoleh nilai minimal B mencapai 70%, tanpa nilai D dan E, mahasiswa dengan kriteria keaktifan pada kategori tinggi dan sangat tinggi mencapai minimal 80%, dan tingkat keterlaksanaan kegiatan dalam pembelajaran berbasis projek dengan pendekatan JAS mencapai 80%. Simpulan yang dapat diambil dari penelitian ini adalah model pembelajaran berbasis projek dengan pendekatan JAS pada perkuliahan Fisiologi Hewan efektif diterapkan

Aggressive Multimodality Treatment for Advanced Rectal Cancer

A case of advanced rectal cancer treated by aggressive local and systemic treatment who has survived more than 7 years from initial recurrence is presented. A 55-year-old woman was diagnosed with advanced lower rectal cancer and underwent a low anterior resection with complete removal of all regional lymph nodes and total mesorectal excision. The tumor was diagnosed as a moderately differentiated adenocarcinoma, pStage IIIB (T3, N2a, M0). Twenty-six months after the initial surgery, local recurrence in the pelvis was detected by computed tomography, and total pelvic exenteration with distal sacrectomy (TPES) was performed after systemic chemotherapy with a molecular-targeted drug. Six months after the TPES, multiple lung metastases were detected. Consequently, the patient underwent radiofrequency ablation (RFA) and chemotherapy. The disease has since been controlled for 38 months. As volume control is essential for cancer treatment, it may be important to combine appropriate local therapy with systemic therapy to metastatic or recurrent sites in order to achieve much longer disease control

Expansion of CpG Methylation in the SFRP2 Promoter Region during Colorectal Tumorigenesis

Secreted frizzled-related protein 2, (SFRP2) is a Wnt inhibitor whose promoter CpGs were recently found to be methylated at high frequency in colorectal cancers (CRCs). We hypothesized that the pattern of SFRP2 methylation may differ throughout the promoter during progressive tumorigenesis. Using combined bisulfite restriction analysis (COBRA), two methylation-sensitive regions (Regions A and B) of the SFRP2 promoter were investigated in 569 specimens of colorectal tissue:222 CRCs, 103 adenomatous polyps (APs), 208 normal colonic mucosa from CRC patients (N-Cs), and 36 normal colonic mucosa from subjects with no evidence of colorectal neoplasia at colonoscopy (N-Ns). Extensive (including both Regions A and B) and partial (either Region A or B) SFRP2 methylation levels were found in 61.7% and 24.8% of CRCs, 8.7% and 37.9% of APs, 3.9% and 39.9% of N-Cs, and 0% and 30.6% of N-Ns, respectively. Extensive methylation of the SFRP2 promoter was present primarily in CRCs, while partial methylation was common in APs. Whereas APs with the KRAS mutant showed no correlation to any pattern of SFRP2 methylation, extensive methylation of the SFRP2 promoter was significantly associated with KRAS mutant CRCs (p＜.0001), suggesting that genetic alteration in the RAS-RAF pathway might precede the spread of CpG methylation through the SFRP2 promoter, which is observed in over 60% of advanced colorectal tumors

A successful rechallenge with cetuximab for a case with metastatic rectal cancer

　A 55-year-old man who had been diagnosed with rectal cancer with multiple liver metastases and lymph node metastases on colonoscopy and computed tomography (CT) was referred to Okayama University Hospital for treatment. Based on the diagnosis of non-curative rectal cancer, we planned to perform systematic chemotherapy after surgical resection. We performed a low anterior resection of a 36×35 mm upper rectal moderately differentiated adenocarcinoma with wil-type KRAS. After the resection, a FOLFIRI regimen with cetuximab was given as the first-line chemotherapy. Although metastatic lesions in the liver showed shrinkage, we decided to switch regimens because of intolerable adverse events. A modified FOLFOX6 regimen with bevacizumab was administered as the second-line treatment. There were no signs of disease progression until eight months later, when positron emission tomography (PET)/CT scans revealed that the new metastatic lesions appeared. As the third-line treatment, an irinotecan with cetuximab regimen was administered, leading to a good response for over 12 months. 　We experienced a successful rechallenge with cetuximab for a case with metastatic rectal cancer. For patients with wild-type KRAS colorectal cancer, rechallenge with cetuximab-based chemotherapy can be an effective therapeutic option

Fluorescence virus-guided capturing system of human colorectal circulating tumour cells for non-invasive companion diagnostics

Background Molecular-based companion diagnostic tests are being used with increasing frequency to predict their clinical response to various drugs, particularly for molecularly targeted drugs. However, invasive procedures are typically required to obtain tissues for this analysis. Circulating tumour cells (CTCs) are novel biomarkers that can be used for the prediction of disease progression and are also important surrogate sources of cancer cells. Because current CTC detection strategies mainly depend on epithelial cell-surface markers, the presence of heterogeneous populations of CTCs with epithelial and/or mesenchymal characteristics may pose obstacles to the detection of CTCs. Methods We developed a new approach to capture live CTCs among millions of peripheral blood leukocytes using a green fluorescent protein (GFP)-expressing attenuated adenovirus, in which the telomerase promoter regulates viral replication (OBP-401, TelomeScan). Results Our biological capturing system can image epithelial and mesenchymal tumour cells with telomerase activities as GFP-positive cells. After sorting, direct sequencing or mutation-specific PCR can precisely detect different mutations in KRAS, BRAF and KIT genes in epithelial, mesenchymal or epithelial–mesenchymal transition-induced CTCs, and in clinical blood samples from patients with colorectal cancer. Conclusions This fluorescence virus-guided viable CTC capturing method provides a non-invasive alternative to tissue biopsy or surgical resection of primary tumours for companion diagnostics

A case of metastatic cecal cancer with mutation in the BRAF oncogene and poor survival

　A 79-year-old woman visited a previous hospital with a complaint of general fatigue. The patient was diagnosed with cecal cancer with multiple liver metastases and lymph node metastases on colonoscopy, abdominal ultrasonography and CT scan, and was referred to our division for treatment. Based on the diagnosis of non-curative colonic cancer, we planned to perform systematic chemotherapy after local surgical treatment. We performed an ileocecal resection, and the specimen showed poorly differentiated adenocarcinoma with mutation in the BRAF oncogene. After the surgical treatment, the tumor grew rapidly and the patient died from cancer on the 19th postoperative day without having the opportunity to undergo chemotherapy. 　Multiple genetic and epigenetic alterations in oncogenes and tumor suppressor genes are involved in the process of colorectal carcinogenesis. Some of the alterations have been identified as predictive and prognostic biomarkers. A mutation in the BRAF oncogene was reported to be associated with a very unfavorable prognosis in colorectal cancers. Some of the cases with rapid progression are suggested to have the BRAF oncogene mutation. According to our experience, chemotherapy before surgical treatment might improve the prognosis of cases with the BRAF mutation

Concordance of acquired mutations between metastatic lesions and liquid biopsy in metastatic colorectal cancer

Aim: To evaluate whether PCR-reverse sequence-specific oligonucleotide can examine the concordance between liquid biopsy and metastatic lesions with acquired resistance. Materials & methods: We examined acquired mutations in chemoresistant lesions and blood obtained from four patients with RAS wildtype metastatic colorectal cancer who underwent treatment with anti-epidermal growth factor receptor antibodies. Results: In one patient, metastatic lesions harbored diverse acquired mutations in KRAS in all seven metastases; the two acquired mutations were detectable in blood collected after the patient acquired resistance. None of the other patients exhibited liquid biopsy mutations, except one, with a BRAF mutation confirmed in primary tumor and peritoneal dissemination. Conclusion: Liquid biopsy based on PCR-reverse sequence-specific oligonucleotide is a successful procedure for capturing acquired mutations with precise information on the RAS mutational spectrum