CDCA2 promotes proliferation and migration of melanoma by upregulating CCAD1.

OBJECTIVE: This study aims at investigating the functional role of CDCA2 (cell division cycle associated 2) in enhancing proliferative and migratory abilities in melanoma by upregulating CCAD1, thus aggravating the progression of melanoma. PATIENTS AND METHODS: CDCA2 levels in melanoma tissues and cell lines were determined by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) and Western blot. Regulatory effects of CDCA2 on proliferative and migratory abilities in melanoma cells were assessed by Cell Counting Kit-8 (CCK-8), 5-Ethynyl-2'-deoxyuridine (EdU), and wound healing assay, respectively. At last, rescue experiments were conducted to explore the involvement of CCAD1 in CDCA2-regulated progression of melanoma. RESULTS: CDCA2 was upregulated in melanoma tissues, especially in those with metastasis. Identically, in vitro level of CDCA2 was upregulated in melanoma cell lines. The knockdown of CDCA2 in A375 and sk-mel-110 cells inhibited the proliferative and migratory abilities. The overexpression of CCAD1 could partially abolish the inhibitory effects of silenced CDCA2 on proliferative and migratory abilities in melanoma. CONCLUSIONS: CDCA2 stimulates proliferative and migratory abilities in melanoma cells by upregulating CCAD1, thus aggravating the malignant progression of melanoma.

Role of asparagine-linked oligosaccharides in the function of the rat PTH/PTHrP receptor.

The receptor for parathyroid hormone (PTH) and PTH-related peptide (PTHrP) is a G-protein-coupled receptor with four potential sites for N-linked glycosylation. The contribution of the oligosaccharide moieties to cell surface expression, ligand binding, and signal transduction was investigated. Site-directed mutagenesis of the rat PTH/PTHrP receptor cDNA was performed at single or combination of the four potential glycosylation sites to determine the effect of the putative carbohydrate chains on the activities of the receptor. The results revealed that all four potential N-glycosylation sites in the PTH/PTHrP receptor are glycosylated. Receptors missing a single or multiple glycosylation consensus but with at least one intact glycosylation site expressed sufficiently and functioned normally. In contrast, the nonglycosylated receptor, in which all four glycosylation sites were mutated, is deficient in these functions. These data indicate important roles for N-linked glycosylation in PTH/PTHrP receptor functions.

Direct mapping of an agonist-binding domain within the parathyroid hormone/parathyroid hormone-related protein receptor by photoaffinity crosslinking.

Parathyroid hormone (PTH) and PTH-related protein (PTHrP) are calciotropic hormones interacting with a shared seven-transmembrane domain G protein-coupled receptor, which is located predominantly in bone and kidney. To map the interface of the bimolecular interaction between hormone and receptor, we designed and radioiodinated a bioactive, photoreactive PTH agonist, (125)I-[Nle(8,18),Lys13(epsilon-p-(3-I-Bz)Bz),L-2-Nal(23),Arg(26,2 7),Tyr34] bPTH-(1-34)NH2 ((125)I-all-R-K13). This ligand contains a photoreactive benzophenone moiety attached to the side chain of Lys13. All other lysyl residues are substituted by argynyls. The analog photocrosslinks specifically to the recombinant hPTH/PTHrP receptor stably transfected into human embryonic kidney cells (HEK-293/C-21 cells, approximately 400,000 receptors per cell), generating a diffuse approximately 87-kDa band on SDS/PAGE autoradiography. To identify the "contact domain" within the hPTH/PTHrP receptor involved in binding of (125)I-all-R-K13, the radiolabeled band containing the ligand-receptor conjugate was subjected to chemical and enzymatic cleavage. Two independent pathways of sequential digestion were used: Route A, lysyl endopeptidase C, then endo-N-glycosidase F, followed by cyanogen bromide; Route B, cyanogen bromide followed by endo-N-glycosydase F. The identified domain is in contact with position 13 in (125)I-all-R-K13 and corresponds to amino acids 173-189 of the hPTH/PTHrP receptor, located at the C-terminal region of the N-terminal extracellular domain.

Free flap coverage for knee salvage.

Soft-tissue reconstruction using free flaps was reviewed in 11 consecutive knee wounds complicated by joint exposure or adjacent osteomyelitis and unavailability of adequate local flaps. Ten free muscle flaps and one fasciocutaneous flap were used successfully. Eight limbs presented with wide exposure of the knee joint, including 4 patients with wound infection involving the joint. Three other patients suffered osteomyelitis immediately adjacent to the knee. All knees were successfully salvaged with a single free flap operation, except for one knee that required a second free flap after flap necrosis. Follow-up in nine flaps ranged from 7 weeks to 19 months (mean, 12 months). For patients without preexisting conditions affecting the knee, free flap coverage provided excellent return of knee function. The choice of recipient vessels was determined mainly by the region of the knee requiring coverage. The recipient arteries utilized included the distal superficial femoral artery, anterior tibial artery, popliteal artery, and small arteries, which included the saphenous artery and descending branch of the lateral circumflex femoral artery. The use of small arteries and venae comitantes as recipient vessels increased the effective reach of the free flaps and eliminated the need for vein grafts.

Clinical evaluation of a rapid immunochromatographic assay based on the 38 kDa antigen of Mycobacterium tuberculosis on patients with pulmonary tuberculosis in China.

SETTING: A rapid membrane-based antibody assay capable of diagnosing pulmonary tuberculosis within 15 min has been developed using the 38 kDa antigen from Mycobacterium tuberculosis. OBJECTIVE: To determine the specificity and sensitivity of this assay and evaluate its usefulness in a clinical setting. DESIGN: Sera from patients with active pulmonary tuberculosis were obtained from three hospitals in China. The control groups consisted of patients who were diagnosed with lung diseases other than tuberculosis and healthy subjects. RESULTS: Antibody was detected in 54 of 61 (89%) sputum positive patients and 67 of 91 (74%) sputum negative patients who had been clinically diagnosed as having active pulmonary tuberculosis. Five out of 56 (9%) patients with respiratory diseases other than tuberculosis and 1 out of 30 (3%) healthy controls had a positive antibody response. The overall specificity of the assay was 93% and the positive predictive value was 95%. We conclude that this assay is rapid, sensitive and specific and will be a valuable aid in the clinical diagnosis of pulmonary tuberculosis.

Double skin paddle fibular flap for a through-and-through oromandibular defect.

The fibular osteocutaneous free flap has become the most versatile method for reconstructing composite defects of the mandible. The complexity of the reconstruction increases substantially when both intraoral and extraoral defects are present, possibly dictating the need for an additional flap. For the smaller through-and-through defects, a single skin island may be sufficient, but may be limited by inadequate reach. Although not routinely necessary, the skin island may be split completely to enhance the effective reach of the second paddle. One patient report is presented for which a moderately sized through-and-through defect of the mandible was reconstructed using a double skin paddle fibular osteocutaneous free flap.

Detection of pulmonary and extrapulmonary tuberculosis patients with the 38-kilodalton antigen from Mycobacterium tuberculosis in a rapid membrane-based assay.

A rapid membrane-based serologic assay using the 38-kDa antigen from Mycobacterium tuberculosis for the diagnosis of tuberculosis (TB) was evaluated with 201 patients with pulmonary TB, 67 patients with extrapulmonary TB, 79 Mycobacterium bovis BCG-vaccinated healthy controls, and 77 non-TB respiratory patients. The overall sensitivities, specificities, and positive and negative predictive values were, respectively, 92, 92, 84, and 96% for sputum-positive TB patients; 70, 92, 87, and 79% for sputum-negative TB patients; and 76, 92, 80, and 90% for extrapulmonary-TB patients. Only 2% (1 of 44) of the healthy control BCG-vaccinated subjects gave weak positive signals in the assay, indicating that this rapid serological assay is a valuable aid in clinical diagnosis for both pulmonary and extrapulmonary TB.

Long-term sequelae following median sternotomy wound infection and flap reconstruction.

Use of muscle and omental flaps has been shown to provide reliable reconstruction of infected median sternotomy wounds; however, few reports emphasize the long-term sequelae of the complication and its treatment. This study was performed to evaluate the long-term problems, including patient satisfaction and survival rate, in 88 patients with median sternotomy infections treated with muscle or omental flaps. Forty-two patients were available for long-term follow-up by telephone interview, with an average length of follow-up of 42 months. Forty-three percent complained of chronic chest wall pain or discomfort, and 45% complained of sternal instability. After pectoralis major muscle flap reconstruction in 32 patients, 25% complained of upper extremity weakness, and 56% complained of chest contour deformity. Delayed septic costochondritis or osteomyelitis occurred in 8%. Despite these unfavorable consequences, 72% and 83% of patients were satisfied with the cosmesis of the operation and the overall result, respectively. Furthermore, after hospital discharge, these patients seem to enjoy satisfactory longevity. By emphasizing the potential sequelae, further research interest may be stimulated in delineating their causes and in refining techniques of reconstruction.