Midkine inhibitors: application of a simple assay procedure to screening of inhibitory compounds

<p>Abstract</p> <p>Background</p> <p>Midkine is a heparin-binding cytokine and is involved in etiology of various diseases. Thus, midkine inhibitors are expected to be helpful in treatment of many diseases.</p> <p>Methods</p> <p>We developed a simple assay for midkine activity based on midkine-dependent migration of osteblastic cells. Midkine inhibitors were searched as materials that inhibit this midkine activity. To develop peptides that inhibit midkine activity, we constructed models in which C-terminal half of midkine interacted with α₄β₁-integrin. Low molecular weight compounds which are expected to bind to midkine with high affinity were searched by <it>in silico </it>screening with the aid of Presto-X2 program.</p> <p>Results</p> <p>Among peptides in putative binding sites of midkine and the integrin, a peptide derived from β₁-integrin and that derived from the first β sheet of the C-terminal half of midkine significantly inhibited midkine activity. Two low molecular weight compounds found by <it>in silico </it>screening exhibited no toxicity to target cells, but inhibited midkine activity. They are trifluoro compounds: one (PubChem 4603792) is 2-(2,6-dimethylpiperidin-1-yl)-4-thiophen-2-yl-6-(trifluoromethy)pyrimidine, and the other has a related structure.</p> <p>Conclusions</p> <p>The assay procedure is helpful in screening midkine inhibitors. All reagents described here might become mother material to develop clinically effective midkine inhibitors.</p

Midkine as a factor to counteract the deposition of amyloid β-peptide plaques: in vitro analysis and examination in knockout mice

<p>Abstract</p> <p>Background</p> <p>Midkine is a heparin-binding cytokine involved in cell survival and various inflammatory processes. Midkine accumulates in senile plaques of patients with Alzheimer's disease, while it counteracts the cytotoxic effects of amyloid β-peptide and inhibits its oligomerization. The present study was conducted to understand the role of midkine upon plaque formation of amyloid β-peptide.</p> <p>Methods</p> <p>A surface plasmon assay was performed to determine the affinity of midkine for amyloid β-peptide. The deposition of amyloid β-peptide was compared in the brain of wild-type and midkine-deficient mice. An effect of midkine to microglias was examined by cell migration assay.</p> <p>Results</p> <p>Midkine bound to amyloid β-peptide with the affinity of 160 nM. The C-terminal half bound to the peptide more strongly than the N-terminal half, and heparin inhibited midkine from binding to the peptide. Pleiotrophin, which has about 50% sequence identity with midkine also bound to amyloid β-peptide. The deposition of amyloid β-peptide plaques in the cortex and hippocampus was more intense in 15-month-old midkine-deficient mice, compared to the corresponding wild-type mice. Midkine promoted migration of microglias in culture.</p> <p>Conclusions</p> <p>These results are consistent with the view that midkine attenuates the deposition of amyloid β-peptide plaques, and thus progression of Alzheimer's disease, by direct binding and also by promoting migration of microglias.</p

Harada\u27s Disease (Vogt-Koyanagi-Harada Syndrome; VKH Syndrome)

We studied the course and outcome of 44 patients with Harada\u27s disease (17 males and 27 females) who were treated in the Department of Ophthalmology of Kawasaki Hospital, an affiliated hospital of Kawasaki Medical School, between April 1980 and March 1990. The age at onset ranged from 14 to 77 years with the mean age±SD being 46.5 ± 15.6 years. Thus the disease was frequent among the elderly. The posterior and optic types of the disease were most frequently found in our patients. As extraocular symptoms, perceptive deafness and an increase in the cell count of the cerebrospinal fluid were frequently observed, and these were important factors in the diagnosis of the disease. Systemic administration of steroids was the main treatment, with immunosuppressive agents being administereted to patiens with the delayed type of the disease. As for the visual prognosis, the final corrected visual acuity was 1.0 in 79% of the patients. As eye complications, cataracts and glaucoma were frequently noted, and various other lesions of the fundus were also observed with increasing age. Patients with the delayed type of the disease accounted for 34.1% of the patients studied

Abnormalities Caused by Carbohydrate Alterations in Iβ6-N-Acetylglucosaminyltransferase-Deficient Mice

Iβ6-N-acetylglucosaminyltransferase (IGnT) catalyzes the branching of poly-N-acetyllactosamine carbohydrate chains. In both humans and mice, three spliced forms of IGnT have been identified, and a common exon is present in all of them. We generated mice deficient in the common exon to understand the physiological function of poly-N-acetyllactosamine branching. IGnT activity was abolished in the stomach, kidney, bone marrow, and cerebellum of the deficient mice, while a low level of the activity persisted in the small intestine. Immunohistochemical analysis confirmed the loss of I antigen from the lung, stomach, and kidney. The deficient mice had reduced spontaneous locomotive activity. The number of peripheral blood lymphocytes was also reduced and renal function decreased in the deficient mice. Furthermore, in aged mice, vacuolization occurred in the kidney, and epidermoid cysts were frequently formed. However, cataracts did not develop earlier in the deficient mice. Decreased levels of lysosomal proteins, LAMP-2 and synaptotagmin VII, were found in the kidney of the deficient mice and correlated with renal abnormalities