Nucleotide sequence of the G protein gene of human respiratory syncytial virus reveals an unusual type of viral membrane protein.

The major surface glycoprotein (G) of human respiratory syncytial (RS) virus has an estimated mature Mr of 84,000-90,000. Among a library of cDNA clones prepared from RS virus mRNAs, we identified clones that hybridized to a message that encoded a Mr 36,000 polypeptide that was specifically immunoprecipitated with anti-G antiserum. The amino acid sequence of the G protein backbone was determined by nucleotide sequence analysis of several of the cDNA clones. It contains a combination of structural features that make it unique among the known viral glycoproteins. The G mRNA is 918 nucleotides long and contains a single major open reading frame that encodes a polypeptide having 298 amino acid residues with a Mr of 32,587, a finding consistent with the Mr 36,000 estimate for the in vitro translation product of the G mRNA. This suggests that greater than 50% of the molecular weight of the mature glycoprotein may be contributed by carbohydrate. Glycosylation of G is largely resistant to tunicamycin, an inhibitor of the attachment of N-linked oligosaccharides, suggesting that the majority of the carbohydrate residues are attached via O-glycosidic bonds. In accordance with this, serine and threonine residues, the acceptor sites for O-linked oligosaccharides, comprise 30.6% of the total amino acid composition. There are also four potential acceptor sites for N-linked oligosaccharides. The amino acid sequence lacks both an NH2-terminal hydrophobic signal sequence and a COOH-terminal hydrophobic region. Instead, a strongly hydrophobic region is located between amino acid residues 38 and 66. This region may serve as both the signal to insert the nascent polypeptide through the membrane and as the membrane anchor site.     

Endoscopic thyroidectomy via bilateral axillo-breast approach (BABA): review of 512 cases in a single institute

Background

Various techniques for endoscopic thyroidectomy have been introduced in the past decade, and the cosmetic superiority of these techniques has been universally acknowledged. We developed the endoscopic thyroidectomy via bilateral axillo-breast approach (BABA) and have performed more than 500 operations. The aims of this study are to analyze the surgical outcomes and to evaluate the effectiveness and safety of BABA endoscopic thyroidectomy.

Patients and methods

Between February 2004 and March 2008, 512 patients with thyroid diseases underwent BABA endoscopic thyroidectomy. The criteria analyzed were clinicopathologic characteristics, types of operation, operation time, tumor–node–metastasis (TNM) stage on the basis of the 7th edition of the American Joint Committee on Cancer (AJCC), results after radioactive ablation therapy, and recurrence of disease in these patients.

Results

Of 512 patients, 397 had a malignant tumor and 115 had benign thyroid disease. Eight patients were diagnosed with Graves’ disease, and nine patients underwent completion thyroidectomy. Three cases were subjected to open thyroidectomy due to uncontrolled bleeding. Mean operation time was 151.2?±?38.1?min for total and near-total thyroidectomy, and 141.7?±?50.1?min for subtotal thyroidectomy and lobectomy. Regarding postoperative complications, transient hypocalcemia occurred in 31.1% of patients and permanent hypoparathyroidism occurred in 4.2% of patients. Transient hoarseness occurred in 20.3% of patients, and permanent vocal cord palsy occurred in 1.7%. Mean hospital stay after operation was 3.34?±?0.8?days (range 3–7?days), and mean follow-up period was 57.1?±?17.6?months (range 38.5–71.7?months). There were eight cases of recurrent thyroid carcinoma, and no mortality has occurred up to the present time.

Conclusions

Endoscopic thyroidectomy via bilateral axillo-breast approach is a safe and effective method that gives good surgical completeness, a low rate of postoperative complications and recurrence, and an excellent cosmetic result. Therefore, this method is a good choice for patients with surgical thyroid diseases.     

Critical‐size defect induces unicompartmental osteoarthritis in a stable ovine knee

Animal models simulating osteoarthritis are frequently associated with irreversible changes in biomechanics. Although these models successfully induce osteoarthritis, results of experimental repair procedures are impaired by biomechanical problems. The aim of this study was to define the critical size of a chondral lesion to induce unicompartmental osteoarthritis in a stable joint. Sixteen sheep were randomly divided into four treatment groups. A cartilage defect (7‐ or 14‐mm diameter) was created in the weight‐bearing zone of the medial femoral condyle. The sheep were mobilized for 6 or 12 weeks. Osteoarthritis was determined by gross assessment, India ink staining, histology (Mankin score), and analysis of COMP in the serum. In the 6‐week group, only minor osteoarthritis was registered for either defect size. After 12 weeks, the 14‐mm defect induced minor osteoarthritis at the femoral condyle and caused significant degenerative changes at the tibial articular cartilage and the meniscus. The 7‐mm defect created focal unicompartmental osteoarthritis at the medial femoral condyle and minor degenerative changes at the corresponding tibia. A 7‐mm full‐thickness chondral defect with a weight‐bearing regimen of 12 weeks induced local osteoarthritis at the medial compartment in an otherwise stable joint as aimed. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 30:214–220, 2012     

Deleterious effects of discography radiocontrast solution on human annulus cell in vitro: changes in cell viability,proliferation, and apoptosis in exposed cells

Background contextCarragee et al. have recently shown that modern discography injections are associated with subsequent acceleration of disc degeneration, herniation, and loss of disc height. Although needle puncture and pressurization are known trauma events that can create disc degeneration in animal models, another likely culprit in clinical discography–associated degeneration is a direct effect of the contrast agent itself on disc cells.PurposeTo test the hypothesis that discography contrast solution would have a deleterious effect on human annulus cells in vitro.Study designAn in vitro study using cultured human annulus cells to assay cell death, cell proliferation, and apoptosis.Patient sampleAnnulus cells from eight surgical disc specimens were evaluated (two Thompson Grade III discs and six Grade IV discs) for cell death and proliferation, and an additional five cultures were tested for apoptosis.Outcome measuresThe proportion of dead and live cells, cell proliferation, and the proportion of apoptotic cells in control and experimental groups.MethodsAfter internal review board approval, experimental design used two sets of controls: untreated cells under our normal culture conditions (control) and a set with added glucose to adjust the osmolality to match respective Isovue radiocontrast solution treatments (glucose controls) using a freezing point osmometer. Treated cells received Isovue 200 (iopamidol, Isovue-M 200; Bracco Diagnostics, Inc., Princeton, NJ, USA) at 12.5, 25, 50, or 100 mg/mL. Twenty thousand cells/well were seeded in triplicate in 24 well plates, control or test media added, and incubated for 24 hours. At termination, dead cells were identified with trypan blue staining and percentage dead cells determined. Cells were also tested to determine the percentage of apoptotic cells after 50 or 100 mg/mL Isovue exposures. Proliferation assays used standard plate reader methods. Statistical analysis used repeated measures analysis of variance with SAS software (version 9.2; SAS Institute, Inc., Cary, NC, USA).ResultsAnalysis of cell death showed a significant increase in the percentage of dead cells with increasing Isovue concentrations compared with control cells (p=.018–.0008). Cell proliferation analyses showed significantly reduced division in Isovue-treated cells (p=.004), and apoptosis assays revealed a significantly higher proportion of apoptotic cells in cells exposed to 50 and 100 mg/mL Isovue (p=.016 and .0003, respectively).ConclusionsDiscography is used extensively in the evaluation of low back pain. Because the lifetime prevalence of disc degeneration and low back pain is high (80% in the general population), many patients may undergo this procedure. Data presented here show that cells exposed in vitro to a radiocontrast agent with adjustments for osmolality have significantly reduced proliferation, increased cell death, and increased programmed cell death (apoptosis). In light of the well-recognized age- and degeneration-related decrease in disc cell numbers, it is possible that radiocontrast exposure may be contributing significantly to disc cell loss with subsequent progression of disc degeneration. Findings presented here provide a plausible cell-based explanation for the previously reported disc degeneration in patients receiving discography contrast solutions.     

Prospective, Randomized, Multi-Center Trial of Antibody Induction Therapy in Simultaneous Pancreas-Kidney Transplantation

A randomized, multicenter, prospective study was conducted at 18 pancreas transplant centers in the United States to determine the role of induction therapy in simultaneous pancreas-kidney (SPK) transplantation. One hundred and 74 recipients were enrolled: 87 recipients each in the induction and noninduction treatment arms. Maintenance immunosuppression consisted of tacrolimus, mycophenolate mofetil, and corticosteroids. There were no statistically significant differences between treatment groups for patient, kidney, and pancreas graft survival at 1-year. The 1-year cumulative incidence of any treated biopsy-confirmed or presumptive rejection episodes (kidney or pancreas) in the induction and noninduction treatment arms was 24.6% and 31.2% (p = 0.28), respectively. The 1-year cumulative incidence of biopsy-confirmed, treated, acute kidney allograft rejection in the induction and noninduction treatment arms was 13.1% and 23.0% (p = 0.08), respectively. Biopsy-confirmed kidney allograft rejection occurred later post-transplant and appeared to be less severe among recipients that received induction therapy. The highest rate of Cytomegalovirus (CMV) viremia/syndrome was observed in the subgroup of recipients who received T-cell depleting antibody induction and received organs from CMV serologically positive donors. Decisions regarding the routine use of induction therapy in SPK transplantation must take into consideration its differential effects on risk of rejection and infection.     

Extended rat-ear flap model: a new rodent model for studying the effects of vessel supercharging on flap viability

A new extended rat-ear flap model, with both an axial and a random component, is described. The flap is based on an axial supply by the posterior auricular artery and the posterior facial vein. The random portion, consisting of the rat dorsum, is capable of being supercharged at two separate sites-in the scapular and pelvic regions. There are several advantages to this composite flap. It is a combined axial and random flap. When used as a free flap, the viability of the axial portion serves as an indicator for anastomotic patency. The random portion allows for the investigation of the effects of pharmaceutical manipulation or surgical intervention, e.g., flap supercharging. The results indicate that the axial supply alone can cover approximately 50 percent of the extended rat-ear flap. Moreover, adding supercharging perforators to the random portion significantly increases the area of flap survival. Of interest, an axial vascular supply, coupled with more distal dorsal perforators (pelvic) than proximal (scapular) perforators, may increase survival for the so-called "watershed" area in the middle of the random portion of the flap. Additionally, this study also investigated the relative importance of arterial supply vs. venous drainage, using the extended rat-ear flap model. The flap was either supercharged with both the perforators of the scapular and pelvic arteries, or both scapular and pelvic veins. The results of the study suggests that augmenting venous drainage provides statistically significant improvement (87 percent vs. 51.6 percent) in increasing flap survival, when compared to augmenting the arterial supply. Arterial supercharging provided no improvement in flap survival, when compared to no supercharging (axial vessels + arterial supercharging, 51.6 percent vs. axial vessels alone, 49.9 percent). The results also suggest that providing adequate venous outflow is more important than providing additional arterial blood, and that impaired venous outflow may contribute to some cases of flap failure. However, it should be kept in mind that the best flap survival occurs with both arterial and and venous supercharging.     

Acute Inflammatory Syndrome Induced by Mycophenolate Mofetil in a Patient Following Kidney Transplantation

Mycophenolate mofetil (MMF) is increasingly used to prevent acute and chronic rejection following kidney transplantation and in autoimmune diseases. Here, we report on a patient after kidney transplantation, who developed an acute inflammatory syndrome characterized by fever and oligoarthritis within 1 week after increasing the MMF dosage. MMF was discontinued resulting in a complete resolution of the syndrome within 48 h. We demonstrated in vitro that the patient's phorbol myristate acetate (PMA-) and formyl Met-Leu-Phe (fMLP-) stimulated polymorphonuclear neutrophils (PMNs) developed increased oxidative burst when incubated with MMF. This report demonstrates that MMF can also induce acute inflammatory syndrome in patients following kidney transplantation and that this syndrome might be due to a paradox, pro-inflammatory reaction of PMNs.     

Comparison of Bilateral Axillo-Breast Approach Robotic Thyroidectomy with Open Thyroidectomy for Graves’ Disease

Background

There is an ongoing debate about whether robotic thyroidectomy (RT) is appropriate for Graves’ disease. The aim of this study was to compare the safety of bilateral axillo-breast approach (BABA) RT with that of open thyroidectomy (OT) in patients with Graves’ disease.

Methods

From January 2008 to June 2014, 189 (44 BABA RT and 145 OT) patients underwent total thyroidectomy for Graves’ disease. Recurrence of Graves’ disease, intraoperative blood loss, hospital stay, and complication rates including recurrent laryngeal nerve (RLN) palsy and hypoparathyroidism were analyzed between BABA RT and OT groups, after propensity score matching according to age, gender, body mass index, surgical indication, the extent of operation, excised thyroid weight, and follow-up period.

Results

No patient experienced recurrence of Graves’ disease after median follow-up of 35.0 months. Intraoperative blood loss (151.8 ± 165.4 mL vs. 134.5 ± 75.4 mL; p = 0.534) and hospital stay (3.4 ± 0.7 day vs. 3.3 ± 0.7 day; p = 0.564) were not different between BABA RT and OT groups. Complication rates including transient RLN palsy (11.4 vs. 11.4 %; p = 1.000), transient hypoparathyroidism (18.2 vs. 20.5 %; p = 0.787), permanent RLN palsy (0 vs. 2.3 %; p = 0.315), and permanent hypoparathyroidism (2.3 vs. 2.3 %; p = 1.000) were also comparable between groups.

Conclusions

BABA RT for Graves’ disease showed comparable surgical completeness and complications to conventional OT. BABA RT can be recommended as an alternative surgical option for patients with Graves’ disease who are concerned about cosmesis.

     

Predictive value of seven preoperative prognostic scoring systems for spinal metastases

Predicting prognosis is the key factor in selecting the proper treatment modality for patients with spinal metastases. Therefore, various assessment systems have been designed in order to provide a basis for deciding the course of treatment. Such systems have been proposed by Tokuhashi, Sioutos, Tomita, Van der Linden, and Bauer. The scores differ greatly in the kind of parameters assessed. The aim of this study was to evaluate the prognostic value of each score. Eight parameters were assessed for 69 patients (37 male, 32 female): location, general condition, number of extraspinal bone metastases, number of spinal metastases, visceral metastases, primary tumour, severity of spinal cord palsy, and pathological fracture. Scores according to Tokuhashi (original and revised), Sioutos, Tomita, Van der Linden, and Bauer were assessed as well as a modified Bauer score without scoring for pathologic fracture. Nineteen patients were still alive as of September 2006 with a minimum follow-up of 12 months. All other patients died after a mean period of 17 months after operation. The mean overall survival period was only 3 months for lung cancer, followed by prostate (7 months), kidney (23 months), breast (35 months), and multiple myeloma (51 months). At univariate survival analysis, primary tumour and visceral metastases were significant parameters, while Karnofsky score was only significant in the group including myeloma patients. In multivariate analysis of all seven parameters assessed, primary tumour and visceral metastases were the only significant parameters. Of all seven scoring systems, the original Bauer score and a Bauer score without scoring for pathologic fracture had the best association with survival (P < 0.001). The data of the present study emphasize that the original Bauer score and a modified Bauer score without scoring for pathologic fracture seem to be practicable and highly predictive preoperative scoring systems for patients with spinal metastases. However, decision for or against surgery should never be based alone on a prognostic score but should take symptoms like pain or neurological compromise into account. A reviewer’s comment on this original article is available at doi:.     

Prostaglandin E1 and misoprostol increase epidermal growth factor production in 3D-cultured human annulus cells

Background contextEpidermal growth factor (EGF) is a peptide known to modulate a number of cellular responses including embryogenesis, cell proliferation, and cell survival. Little is known about EGF and its regulation in human annulus cells. Previous work has identified EGF and its receptor in control outer annulus disc tissue, but not in herniated tissue.PurposeTo determine if human annulus cells express EGF in vitro, to determine if the epidermal growth factor-receptor (EGF-r) was expressed in vivo and in vitro in disc cells, to test the effect of EGF on annulus cell proliferation and proteoglycan production in vitro, and to test the effect of prostaglandin E1 (PGE1) and misoprostol on disc cell production of EGF in vitro.Study design/settingStudies were approved by the authors' Human Subjects Institutional review Board. Human disc tissue was used for immunocytochemistry, and human annulus cells were tested in vitro.Patient sampleThirty-four disc specimens were used for studies of proteoglycan production, cell proliferation, and EGF production in vitro. An additional nine discs were used for EGF-r immunolocalization.MethodsDisc tissue was used for immunocytochemical studies for the localization of EGF-r and as a source for cultured annulus cells. Monolayer culture was used to test proliferation responses to 0, 25, 50, or 75 ng/mL EGF over a 2-day culture period. Three-dimensional (3D) culture in a collagen sponge was used to test 100,000 cells cultured in a paired experimental design over 14 days for production of EGF and proteoglycans. Cells were exposed to control conditions, or to either misoprostol at 8 ng/mL or PGE1 at 10^?7 M. Conditioned media was harvested and assayed using an enzyme-linked immunosorbent assay (ELISA) assay with the Human Protein Cytokine Antibody Array I kit. Replicate EGF relative intensity values were averaged and normalized to controls assayed on the same membrane. 3D-cultured cells were also used to confirm EGF gene expression using microarray analysis. Standard statistical methods were used to analyze results.ResultsMicroarray analysis of mRNA from annulus cells in 3D culture confirmed expression of EGF, and immunocytochemistry verified the presence of EGF-r in vitro and in vivo. PGE1, at a dose of 10^?7 M, and misoprostol (a synthetic PGE1 analog) at a dose of 8 ng/mL, both significantly increased EGF levels in annulus cells cultured in 3D compared with control levels (p=.031 and .034, respectively). No significant difference, however, was seen in cell proliferation or in total sulfated proteoglycan production in EGF-exposed annulus cells.ConclusionsData showed that EGF is expressed and produced by annulus cells in vivo and in 3D culture, with significantly greater in vitro EGF produced in the presence of PGE1 or the PGE1 analog misoprostol. Misoprostol, developed for prevention/treatment of nonsteroidal anti-inflammatory-induced gastropathy, has now been reported to have some interesting anabolic effects stimulating osteoblasts during fracture healing and during ovariectomy in animal models. Exogenous EGF did not increase cell proliferation in monolayer, or total production of proteoglycans in 3D culture. Additional work is needed to further delineate the role of EGF in the human disc.