A Small Compound, HYGIC, Promotes Hypocotyl Growth Through Ectopic Ethylene Response.

Plant seedlings adjust the growth of the hypocotyl in response to surrounding environmental changes. Genetic studies have revealed key players and pathways in hypocotyl growth, such as phytohormones and light signaling. However, because of genetic redundancy in the genome, it is expected that not-yet-revealed mechanisms can be elucidated through approaches different from genetic ones. Here, we identified a small compound, HYGIC (HG), that simultaneously induces hypocotyl elongation and thickening, accompanied by increased nuclear size and enlargement of cortex cells. HG-induced hypocotyl growth required the ethylene signaling pathway activated by endogenous ethylene, involving CONSTITUTIVE PHOTOMORPHOGENIC 1, ETHYLENE INSENSITIVE 2 (EIN2) and redundant transcription factors for ethylene responses, ETHYLENE INSENSITIVE 3 (EIN3) and EIN3 LIKE 1. By using EBS:GUS, a transcriptional reporter of ethylene responses based on an EIN3-binding-cis-element, we found that HG treatment ectopically activates ethylene responses at the epidermis and cortex of the hypocotyl. RNA-seq and subsequent gene ontology analysis revealed that a significant number of HG-induced genes are related to responses to hypoxia. Indeed, submergence, a representative environment where the hypoxia response is induced in nature, promoted ethylene-signaling-dependent hypocotyl elongation and thickening accompanied by ethylene responses at the epidermis and cortex, which resembled the HG treatment. Collectively, the identification and analysis of HG revealed that ectopic responsiveness to ethylene promotes hypocotyl growth, and this mechanism is activated under submergence.

Urinary tract bleeding from a urethral caruncle mimicking genital tract bleeding.

A 69-year-old Japanese woman with a post-hysterectomy status came to our primary care clinic. She presented with vaginal bleeding for the past 3 days which had developed after defecation. There was a palpable mass measuring approximately 2 cm on pelvic exam; however, heavy bleeding prevented in-depth observation. CT and MRI scans revealed that the mass was inside the urethral meatus and not in the vagina. She underwent surgical resection of the urethral tumour, and the pathological report showed no malignancy. A final diagnosis of urethral caruncle was made. Vaginal bleeding is commonly encountered in the primary care practice and is usually attributed to gynaecological diseases. However, patients and physicians may falsely regard urinary or gastrointestinal tract bleeding as one involving the genital tract. We present a case wherein vaginal bleeding was initially considered but was later identified to be due to a urethral caruncle.

Polygenic burdens on cell-specific pathways underlie the risk of rheumatoid arthritis.

Recent evidence suggests that a substantial portion of complex disease risk alleles modify gene expression in a cell-specific manner. To identify candidate causal genes and biological pathways of immune-related complex diseases, we conducted expression quantitative trait loci (eQTL) analysis on five subsets of immune cells (CD4+ T cells, CD8+ T cells, B cells, natural killer (NK) cells and monocytes) and unfractionated peripheral blood from 105 healthy Japanese volunteers. We developed a three-step analytical pipeline comprising (i) prediction of individual gene expression using our eQTL database and public epigenomic data, (ii) gene-level association analysis and (iii) prediction of cell-specific pathway activity by integrating the direction of eQTL effects. By applying this pipeline to rheumatoid arthritis data sets, we identified candidate causal genes and a cytokine pathway (upregulation of tumor necrosis factor (TNF) in CD4+ T cells). Our approach is an efficient way to characterize the polygenic contributions and potential biological mechanisms of complex diseases.

Personal status of general health checkups and medical expenditure: A large-scale community-based retrospective cohort study.

BACKGROUND: We sought to clarify the association between the personal utilization of general health checkups (GHCs) and medical expenditures (MEs) in a middle-aged Japanese population. METHODS: A retrospective cohort study was conducted. Subjects were 33,417 residents (15,819 males and 17,598 females) aged 48 years or older in 2010 who were invited to undergo GHCs every year. Official records on GHCs from 2002 to 2007 and MEs from 2008 to 2010 were provided by Soka City, Saitama Prefecture, Japan. The utilization of GHCs was divided into zero times (non-utilizers), 1-3 times (low-frequency utilizers), and 4-6 times (high-frequency utilizers). Tweedie distributions in the generalized linear model were used to analyze the association between MEs and the subgroups of GHC utilization after adjustment for age and sex. RESULTS: Of the 33,417 subjects, 20,578 (61.6%) were non-utilizers, 5,777 (17.3%) were low-frequency utilizers, and 7,062 (21.1%) were high-frequency utilizers, based on the attendance to GHCs from 2002 to 2007. Compared with the non-utilizers, the high-frequency utilizers showed significantly higher outpatient MEs (JPY394,700 vs. JPY373,100). The low- and high-frequency utilizers showed significantly lower inpatient MEs (JPY224,000 and JPY181,500 vs. JPY309,300) and total MEs (JPY610,600 and JPY580,700 vs. JPY689,600) than the non-utilizers based on the pooled data from 2008 to 2010. CONCLUSIONS: This study suggests that the outpatient MEs rise when annual GHCs are increasingly attended (not including the GHC cost), but inpatient and total MEs are lower. To reduce MEs, increasing the rates of attendance at GHCs by the general public may be important.

[The influence of counseling for patients with cancer on their discharge from the palliative care support department of the community health care service of Minoh City Hospital].

Counseling for patients with cancer by a certified nurse in palliative care began in April 2011 in Minoh City Hospital. Counseling was provided immediately after a patient was informed by the treating physician of a primary diagnosis of cancer, a metastatic recurrence, or a decision to terminate cancer therapy. We examined the patient's support system after the counseling ended. The number of patients receiving end-of-life support with home or hospital care rapidly increased from 118 prior to the program's beginning to 186. The number of patients counseled was comparable to the rapid increase in their number(n=68). New cases in the outpatient department comprised 59% of all patients, of which, 45% began supportive counseling, with 43%of them ultimately returning home. Of the new cases receiving counseling in the hospital, 34%eventually returned home after discharge, and the highest percentage of discharges were to a palliative care unit or hospice program (48%). The initiation of counseling in the outpatient department allowed us to provide sufficient time to make decisions about appropriate places for end-of-life care. Cooperation with the patients' physicians was necessary to provide counseling from the outpatient department. Our findings suggest the importance of sharing the patients' medical and social information among the staff when necessary.

Establishment of a Strategy for the Discovery and Verification of Low-Abundance Biomarker Peptides in Plasma Using Two Types of Stable-Isotope Tags.

Serum and plasma contain thousands of different proteins and peptides, which can provide valuable information about the numerous processes that take place within the body. However, detailed analysis of proteins and peptides in serum and plasma remains challenging due to the presence of many high-abundance proteins, the large dynamic range of protein and peptide concentrations, the extensive complexity caused by posttranslational modifications, and considerable individual variability. In particular, detailed analysis and identification of native peptides is extremely difficult due to the tremendous variety of cleavage possibilities and posttranslational modifications, which results in extremely high complexity. Therefore, widely ranging searches based on peptide identification are difficult. Herein, we describe the highly accurate and sensitive quantitative analysis of over 2,500 peptides with the concentration limit of about 10 pM. The strategy combined isobaric tag labeling, amine-reactive 6-plex tandem mass tag labeling, and a modified differential solubilization method for high-yield peptide extraction [Saito, T. et al. J. Electrophoresis 2013 57: 1-9]. Using this strategy, we quantitatively analyzed six pooled plasma samples (three pre-surgery and three post-surgery) to discover potential candidate biomarker peptides of renal cell carcinoma. The concentrations of 27 peptides were found to be altered following surgery. A preliminary validation study was conducted using about 80 plasma samples to demonstrate the possibility that even unidentified potential candidate biomarker peptides can be verified using the isotope tag/dimethyl labeling method. We also discuss technical consideration and potential of this strategy for facilitating native peptide research.

[Palliative care during chemotherapy for advanced colon cancers].

BACKGROUND: The median survival time following chemotherapy for unresectable metastatic colorectal cancer (mCRC) is approximately 2 years. Although palliative care during the chemotherapy period is very important, it has not been reported in detail. PATIENTS AND METHODS: Information on the palliative care of 110 patients with Stage IV mCRC, who were treated from September 2007 to March 2011, was retrospectively examined. RESULTS: Following an explanation of their recurrence or metastases of mCRC, all the patients received mental care from nurses or psychiatrists. They also needed care to prevent the side effects of chemotherapy. Some patients experienced pain associated with tumor growth. Thus, they required NSAIDs or opioids to reduce the cancer-related pain. After they could not be taken chemotherapy, 87.5% of these patients consulted medical social workers to discuss where they would live. CONCLUSIONS: The patients required palliative care depending on the duration of chemotherapy for mCRC. Thus, we believe that palliative care is an important part of treatment for advanced cancer.

N-Glycosylation of laminin-332 regulates its biological functions. A novel function of the bisecting GlcNAc.

Laminin-332 (Lm332) is a large heterotrimeric glycoprotein that has been identified as a scattering factor, a regulator of cancer invasion as well as a prominent basement membrane component of the skin. Past studies have identified the functional domains of Lm332 and revealed the relationships between its activities and the processing of its subunits. However, there is little information available concerning the effects of N-glycosylation on Lm332 activities. In some cancer cells, an increase of beta1,6-GlcNAc catalyzed by N-acetylglucosaminyltransferase V (GnT-V) is related to the promotion of cancer cell motility. By contrast, bisecting GlcNAc catalyzed by N-acetylglucosaminyltransferase III (GnT-III) suppresses the further processing with branching enzymes, such as GnT-V, and the elongation of N-glycans. To examine the effects of those N-glycosylations to Lm332 on its activities, we purified Lm332s from the conditioned media of GnT-III- and GnT-V-overexpressing MKN45 cells. Lectin blotting and mass spectrometry analyses revealed that N-glycans containing the bisecting GlcNAc and beta1,6-GlcNAc structures were strongly expressed on Lm332 purified from GnT-III-overexpressing (GnT-III-Lm332) and GnT-V-overexpressing (GnT-V-Lm332) cells, respectively. Interestingly, the cell adhesion activity of GnT-III-Lm332 was apparently decreased compared with those of control Lm332 and GnT-V-Lm332. In addition, the introduction of bisecting GlcNAc to Lm332 resulted in a decrease in its cell scattering and migration activities. The weakened activities were most likely derived from the impaired alpha3beta1 integrin clustering and resultant focal adhesion formation. Taken together, our results clearly demonstrate for the first time that N-glycosylation may regulate the biological function of Lm332. This finding could introduce a new therapeutic strategy for cancer.

Sequence specificity and efficiency of protein N-terminal methionine elimination in wheat-embryo cell-free system.

Recent improvements in wheat-embryo cell-free translation resulted in a highly productive system for protein preparation. To clarify N-terminal processing of the cell-free system in a preparative-scale (> mg protein product per ml), 20 mutant variants of maltose-binding protein (MalE), each having a different penultimate residue in the sequence Met-Xaa-Ile-Glu-, and 20 glutathione S-transferase (GST) variants, having Met-Xaa-Pro-Ile-sequence, were designed and synthesized. The MalE and GST proteins were purified by amylose-resin and glutathione columns, respectively, followed by analysis of their N-terminal sequences. These investigations revealed that sequence specificity and efficiency of the N-terminal Met (N-Met) elimination in the cell-free system are similar to those reported from investigations in cellular systems or in the wheat-embryo cell-free protein expression system in analytical scale (approximately 10 microg protein product per ml). Cleavage of the N-Met is basically determined by the penultimate amino acid in the polypeptide sequence. In the case of MalE, the cleavage was efficient when the penultimate residue was Ala, Cys, Gly, Pro, Ser or Thr. But, in the case of GST with Pro as the antepenultimate residue, the efficiency was significantly reduced when the penultimate residue was Gly or Thr. We also confirmed that substitution of the antepenultimate residue in MalE to Pro drastically reduced the efficiency of N-Met cleavage when the penultimate residue was Ala, Gly, Pro, Ser or Thr, indicating inhibitory effects of antepenultimate residue Pro on N-Met elimination. These results clarified sequence-specific functions of the endogenous N-terminal processing machinery in the scaled-up wheat-embryo cell-free translation system.