A randomised controlled trial comparing sugammadex and neostigmine at different depths of neuromuscular blockade in patients undergoing laparoscopic surgery.

Deep neuromuscular blockade during certain surgical procedures may improve operating conditions. Sugammadex can be used to reverse deep neuromuscular blockade without waiting for spontaneous recovery. This randomised study compared recovery times from neuromuscular blockade induced by rocuronium 0.6 mg.kg(-1), using sugammadex 4 mg.kg(-1) administered at 1-2 post-tetanic count (deep blockade) or neostigmine 50 µg.kg(-1) (plus atropine 10 µg.kg(-1)) administered at the re-appearance of the second twitch of a train-of-four stimulation (moderate blockade), in patients undergoing laparoscopic surgery. The primary efficacy variable was the time from the start of sugammadex/neostigmine administration to recovery of the train-of-four ratio to 0.9. Patients receiving sugammadex recovered 3.4 times faster than patients receiving neostigmine (geometric mean (95% CI) recovery times of 2.4 (2.1-2.7) and 8.4 (7.2-9.8) min, respectively, p<0.0001). Moreover, 94% (62/66) of sugammadex-treated patients recovered within 5 min, vs 20% (13/65) of neostigmine-treated patients, despite the difference in the depth of neuromuscular blockade at the time of administration of both drugs. The ability to provide deep neuromuscular blockade throughout the procedure but still permit reversal at the end of surgery may enable improved surgical access and an enhanced visual field.

Subcellular compartmentalization of saccharide moieties in cultured normal and malignant cells.

We studied subcellular localization of saccharide moieties in cultured normal and malignant cells fixed in paraformaldehyde and treated with a nonionic detergent, using lectins specific for various surgar residues as probes in fluorescence microscopy. In normal cells, concanavalin A and Lens culinaris agglutinin, specific for mannose-rich carbohydrate cores in glycoproteins, labeled the endoplasmic reticulum as a wide perinuclear region. Other lectins, on the other hand, stained the Golgi apparatus as a juxtanuclear reticular structure. A similar compartmentalization was also seen in all malignant cells studied, although the Golgi apparatus in these cells was distinctly vesicular in appearance. Our results indicate that saccharide moieties in both normal and malignant cells are similarly compartmentalized, and thus speak in favor of a unidirectional subcellular flow for both membrane and secreted glycoconjugates.

Fibronectin expression is determined by the genotype of the transformed parental cells in heterokaryons between normal and transformed fibroblasts.

The expression of fibronectin, a cell surface-associated transformation-sensitive glycoprotein, was studied in hetero- and homokaryons of normal and SV40-transformed human fibroblasts. In immunofluorescence, fibroblast homokaryons had an intense surface-associated and intracelluar fibronectin fluorescence similar to that of normal fibroblasts. Transformed cells and their homokaryons had a minimal surface-associated and a weak intracellular fibronectin fluorescence. In heterokaryons formed between transformed and normal fibroblasts, the expression of fibronectin fell within 24 h to the level of the transformed cell homokaryons. The change was detectable already at 3 h after fusion and was gene-dose dependent. These results show that the transformed genotype determines fibronectin expression in the heterokaryons.

Expression and distribution of vimentin and keratin filaments in heterokaryons of human fibroblasts and amnion epithelial cells.

The expression of intermediate filaments of the keratin- and the vimentin-type was studied in heterokaryons of human fibroblasts and amnion epithelial cells by immunofluorescence microscopy. Fibroblasts and their homokaryons showed a fibrillar, vimentin-specific fluorescence throughout the cytoplasm but were negative when stained for keratin. Amnion epithelial cells and their homokaryons, on the other hand, showed a keratin-specific fibrillar staining, and only some of them contained also detectable vimentin. When suspended epithelial cells were fused with adherent fibroblasts, keratin fibrils spread within 3 h into the fibroblasts, intermixing with the vimentin fibrils. 1-3 d after fusion, both vimentin and keratin filaments were expressed as typical fibrillar cytoplasmic arrays, and the distribution of keratin in heterokaryons resembled closely that of vimentin. A typical cell-to-cell arrangement of keratin fibrils, seen in cultures of amnion epithelial cells, could also be found between heterokaryons. Treatment of the cultures with vinblastine sulphate induced coiling of the vimentin filaments in both homo- and heterokaryons, whereas the keratin organization was only slightly affected. Our results show that both vimentin and keratin filaments are incorporated into the cytoskeleton of heterokaryons formed between fibroblasts and epithelial cells, and that they behave in the same way as in their parental cells. Both epithelial and fibroblastic characteristics thus appear to the coexpressed in such heterokaryons.

Deletion of the paired alpha 5(IV) and alpha 6(IV) collagen genes in inherited smooth muscle tumors.

The gene encoding alpha 6(IV) collagen, COL4A6, was identified on the human X chromosome in a head-to-head arrangement and within 452 base pairs of the alpha 5(IV) collagen gene, COL4A5. In earlier studies, intragenic deletions of COL4A5 were detected in a subset of patients with Alport syndrome (AS), a hereditary defect of basement membranes. In some families, AS cosegregates with diffuse leiomyomatosis (DL), a benign smooth muscle tumor diathesis. Here it is shown that patients with AS-DL harbor deletions that disrupt both COL4A5 and COL4A6. Thus, type IV collagen may regulate smooth muscle differentiation and morphogenesis.

Short- and long-term mortality and causes of death after reconstruction of cancers of the head and neck with free flaps.

The use of free flaps to reconstruct cancers of the head and neck is accompanied by appreciable postoperative morbidity and high long-term mortality, but the causes of death and the impact of postoperative complications on survival have not been well studied. We have therefore analysed retrospectively the causes of death and survival of 146 such patients operated on between 2008 and 2016 of whom a total of 62 (43%) had died by the end of 2016. The cause of death was the primary disease in 45 of the 62. The median survival of those who died with the primary cancer as the cause of death did not differ from that of those who died of other causes. In a multivariate Cox model indicators of five-year mortality were male sex, low body mass index (BMI), American Society of Anesthesiologists (ASA) grade more than II, and late medical complications. Neither the size of the tumour nor any operative factors were independent risks for five-year mortality. Ten patients died within six months of operation, all of whom had higher postoperative C-reactive protein concentrations than those who survived for more than six months. The cause of death of most patients who died after free flap operations for head and neck cancer was the primary diagnosis. According to these results, patient-related factors (male sex, ASA grade more than II, low BMI, and low albumin concentration) have an important role in long-term survival, which highlights the importance of careful selection of patients for operative treatment.

Complications and outcome after free flap surgery for cancer of the head and neck.

We retrospectively studied 136 patients who had free flap reconstruction for cancer of the head and neck at a single centre (2008-2015) to evaluate complications, assess factors associated with them, and analyse their impact on outcome. Preoperative and perioperative data, and surgical and medical complications were recorded, and the impact of the complications on duration of hospital stay and survival were assessed. A total of 86 (63%) patients had complications. Compared with those who did not, they had a higher rate of alcohol abuse (21/86, compared with 5/50, p=0.039), longer operations (median (IQR) 565 (458-653 compared with 479 (418-556) minutes, p<0.001), and greater intraoperative loss of blood (725 (400-1150) compared with 525 (300-800) ml, p=0.042). Complications were more common in patients who had fibular flaps and T4 disease (22/86 compared with 4/50, p=0.010; 47/80 compared with 16/47, p=0.015, respectively). Those who had complications also stayed in hospital longer (median (IQR) 9 (7-12) compared with 15 (10-21) days, p<0.001). Cumulative mortality was higher in patients with late complications (those that occurred after the fourth postoperative day) (61% compared with 36%, p=0.004). In conclusion, complications in more than half the patients were related to alcohol abuse, a more complicated intraoperative course, and fibular flaps. Complications were associated with a longer hospital stay, and survival was higher in those who did not have late complications than in those who did.

Visual cycle and its metabolic support in gecko photoreceptors.

Photoreceptors of nocturnal geckos are transmuted cones that acquired rod morphological and physiological properties but retained cone-type phototransduction proteins. We have used microspectrophotometry and microfluorometry of solitary isolated green-sensitive photoreceptors of Tokay gecko to study the initial stages of the visual cycle within these cells. These stages are the photolysis of the visual pigment, the reduction of all-trans retinal to all-trans retinol, and the clearance of all-trans retinol from the outer segment (OS) into the interphotoreceptor space. We show that the rates of decay of metaproducts (all-trans retinal release) and retinal-to-retinol reduction are intermediate between those of typical rods and cones. Clearance of retinol from the OS proceeds at a rate that is typical of rods and is greatly accelerated by exposure to interphotoreceptor retinoid-binding protein, IRBP. The rate of retinal release from metaproducts is independent of the position within the OS, while its conversion to retinol is strongly spatially non-uniform, being the fastest at the OS base and slowest at the tip. This spatial gradient of retinol production is abolished by dialysis of saponin-permeabilized OSs with exogenous NADPH or substrates for its production by the hexose monophosphate pathway (NADP+glucose-6-phosphate or 6-phosphogluconate, glucose-6-phosphate alone). Following dialysis by these agents, retinol production is accelerated by several-fold compared to the fastest rates observed in intact cells in standard Ringer solution. We propose that the speed of retinol production is set by the availability of NADPH which in turn depends on ATP supply within the outer segment. We also suggest that principal source of this ATP is from mitochondria located within the ellipsoid region of the inner segment.

The expression of extracellular matrix and cytoskeleton in fused cells.

In the present paper results from our studies on the expression and regulation of the differentiated normal and transformed epithelial and fibroblastic phenotypes are reviewed. The expression of the extracellular matrix- and basement membrane-associated proteins, fibronectin and laminin, and different intermediate filament proteins were studied in different fused cells. Heterokaryons and cytoplasmic hybrids (cybrids) were formed by fusing normal or malignant epithelial cells with normal fibroblasts or malignant glial cells. No differences were observed in the expression of these phenotypic markers between unfused parental cells and the corresponding homokaryons. Thus, the fusion process itself does not cause changes in the expression of these phenotypic markers. In heterokaryons formed after fusion of normal or malignant epithelial cells with normal fibroblasts or malignant glial cells, two or even three different types of intermediate filaments could be co-expressed. Thus, no suppression of the expression of the various intermediate filaments is caused by the non-homologous parental cell. Expression of the extracellular matrix proteins, fibronectin and laminin, on the other hand, could be extinguished by cells and cytoplasts not expressing fibronectin matrix, such as transformed fibroblasts and HeLa cells. These results support the role of transacting regulatory factors controlling the expression of the different extracellular matrix proteins. Alternatively, the results can be explained by changes in the distribution and concentration of the extracellular matrix receptor proteins in the fused cells.

Fluorescent antibodies and lectins stain intracellular structures in fixed cells treated with nonionic detergent.

Nonionic detergent (NP40) treatment of paraformaldehyde-fixed normal and SV40-transformed human fibroblasts resulted in intracellular penetration of two chosen fluorescent antibodies and Concanavalin A (Con A). After the detergent treatment nuclear SV40 T antigen, cytoplasmic fibronectin glycoprotein and Con A binding sites could be visualized in fluorescence microscopy. The lowest NP40 concentration which made fixed cells permeable was 0.05%. The morphology of cells was preserved better by this new method than by conventional fixation methods, such as acetone treatment. In scanning electron microscopy the surface of the fixed NP40-treated cells had only small rugosities and fine pores. The subsurface cytoskeleton especially was well preserved and had a more distinct fine structure. The improved morphology made it possible to detect a similar distribution of fibronectin and Con A binding sites in the perinuclear endoplasmic reticulum regions.

Prognostic value of tumour vascularity in metastatic melanoma and association of blood vessel density with vascular endothelial growth factor expression.

Tumour angiogenesis is essential for tumour growth and metastasis. Several lines of evidence indicate that vascular endothelial growth factor (VEGF) is a major regulator both of physiological and pathological angiogenesis. In this study we assessed the blood vessel density and VEGF expression of 94 melanoma metastases of 70 patients by immunohistochemistry, utilizing antibodies against human platelet endothelial cell adhesion molecule-1 (PECAM-1, CD31) and VEGF. The number of blood vessels ranged from 4 to 131 vessels/high power field (HPF), with a mean value of 32 vessels/HPF (+/-21) and a median of 29 vessels/ HPF. Survival since diagnosis of the primary disease and from the start of chemoimmunotherapy, as well as the disease-free survival period, was significantly shorter in the high vascularity group of patients compared with the low vascularity group (P< 0.05 and P< 0.01, respectively). A high overall expression of VEGF in the metastatic melanoma samples was observed. The degree of VEGF expression appeared to have a strong association with the blood vessel density (P= 0.017). This study demonstrates the clinical role of tumour vascularity in the prognosis of patients with metastatic melanoma. In addition, the strong association between vascularity and VEGF expression suggests a crucial role for this growth factor in the neovascularization of metastatic melanoma.

Atresia of left coronary ostium and left main coronary artery.

An uncommon coronary anomaly incidentally found at autopsy of a 76-year-old woman is reported. There was no history of cardiovascular symptoms or clinical signs of ischemic heart disease. The autopsy disclosed that the patient had no left coronary ostium and that the left main coronary artery was atretic. However, the rest of the left coronary system, including the left anterior descending coronary artery and the left circumflex coronary artery, were intact in their normal positions. Arterial flow to the left coronary system was supplied by a dilated collateral vessel that originated from the right main coronary artery and coursed between the right ventricular infundibulum and the aorta. Our case suggests that atresia of the left coronary ostium and the left main coronary artery, often associated with death at an early age, may also be compatible with symptom-free longevity. Moreover, attention is focused on the nomenclature of single coronary anomalies.

Müllerian adenosarcomalike tumor of the seminal vesicle. A case report with immunohistochemical and ultrastructural observations.

A müllerian adenosarcomalike tumor of the seminal vesicle is described in a 49-year-old man. The tumor occupied the entire right seminal vesicle and adhered to the right lobe of the prostate, in the area adjacent to the seminal vesicle. The tumor was also adherent to the rectum. Microscopically, the cells were seen to invade the prostatic tissue. The tumor consisted of a highly cellular stroma with spindle-shaped pleomorphic cells, suggesting the diagnosis of a low-grade sarcoma. In addition, dilated cystic spaces lined by columnar epithelium were seen. Immunohistochemically, most tumor cells showed positivity for vimentin, desmin, and muscle-specific actin, suggesting smooth-muscle cell differentiation. Furthermore, electron microscopy also demonstrated smooth-muscle differentiation of the tumor cells. The patient has been disease-free for 48 months since undergoing a cystoprostatetectomy.

Exceptional pharmacokinetics of trivalent chromium during occupational exposure to chromium lignosulfonate dust.

The excretion of chromium in the urine of workers exposed to chromium lignosulfonate was studied. The chromium in the dust was in the trivalent (III) oxidation state, and 30% of the particles were less than 5 micron in diameter. Chromium (III) lignosulfonate dust was rapidly absorbed, and a peak of urinary excretion was seen immediately after exposure. No appreciable accumulation of chromium occurred over 3 d, as evaluated by comparison with preshift urinary chromium concentrations. The addition of ethylenediaminetetra-acetate to the urine of exposed persons greatly enhanced the capacity of chromium to traverse a dialysis membrane; the same effect was seen with chromium (III) chloride. It is concluded that chromium (III) lignosulfonate yields chromium (III), which acts pharmacokinetically like water-soluble hexavalent chromium compounds.

Effect of epidermal growth factor on tympanic membranes with chronic perforations: a clinical trial.

Epidermal growth factor is an important modulator of cell growth, and its role in normal wound healing is well documented. Epidermal growth factor receptors have been identified in tympanic membranes of different animals. The ability of epidermal growth factor to promote healing of tympanic membrane perforations has recently been shown in experimental animals. We performed a double-blind, placebo-controlled study of the effect of epidermal growth factor applied locally on the tympanic membrane for 1 week in patients with chronic perforations. Seventeen adult patients took part in the study, eight in the epidermal growth factor group and nine in the placebo group. Three placebo-treated patients were later treated with epidermal growth factor, and five patients received repeated epidermal growth factor treatment. Perforation size was measured as a percentage of the tympanic membrane area before and at least 1 month (mean, 2.6 months) after treatment. One perforation in the placebo group healed completely, but none of the epidermal growth factor-treated perforations closed. Perforations became slightly smaller in both groups (mean decrease, 0.3% and 2.7% for epidermal growth factor and placebo, respectively), but these changes in size were not statistically significant for either group. At otomicroscopy, a proliferation reaction with thickening of the tympanic membrane and pseudomembrane formation at the perforation edge could be seen in some ears. Histologically, a sample from one epidermal growth factor-treated ear demonstrated signs of hypertrophic epithelium when compared with the morphology of a placebo-treated tympanic membrane. The only complications were two mild infections in the placebo group. Hearing remained stable after epidermal growth factor treatment.

Colposcopy referral rate can be reduced by high-risk human papillomavirus triage in the management of recurrent atypical squamous cells of undetermined significance or low-grade squamous intraepithelial lesion cytology in Finland.

The aim of this study was to establish whether a combination of high-risk human papillomavirus (hrHPV) testing and cervical cytology could reduce colposcopy referral among women with atypical squamous cells of undetermined significance (ASCUS) or low-grade squamous intraepithelial lesion cytology. We randomized 598 women in the Helsinki area, Finland into three study groups. Different strategies of hrHPV testing, cytology and colposcopy with biopsy were used; subsequent hrHPV test results and cytological findings were compared with histology. The rates of hrHPV positivity and CIN2+ were compared. In total, 62.5% of all samples were hrHPV-positive. Altogether 45 (12.7%) CIN2 or worse (CIN2+) lesions were found in study groups A and B. Among hrHPV-positive women the rate of CIN2+ was 19.0% (n = 43), in contrast with 1.6% (n = 2) among hrHPV-negative women (relative risk = 12.2, 95% confidence interval [CI] 3.6-81.1, P < 0.001). Among all hrHPV-negative women whose cytological findings were normal or ASCUS, dysplastic lesions were uncommon (n = 4/119, 3.4%), and all were CIN1. If these women had not been referred to colposcopy, the number of colposcopies would have been reduced by 33.6%. We conclude that hrHPV testing combined with repeat cervical cytology had a high negative predictive value in patients with recurrent low-grade cervical cytology. This could reduce the referral rate to colposcopy without jeopardizing patient safety.

Development of a multi-component fiber-reinforced composite implant for load-sharing conditions.

Fiber-reinforced composites (FRC) have the potential for use as load-bearing orthopaedic implants if the high strength and elastic modulus of FRC implant can be matched with local requirements. This study tested the in vivo performance of novel FRC implants made of unidirectional glass fibers (E-glass fibers in Bis-GMA and TEGDMA polymeric matrix). The implant surface was covered with bioactive glass granules. Control implants were made of surface-roughened titanium. Stress-shielding effects of the implants were predicted by finite element modelling (FEM). Surgical stabilization of bone metastasis in the subtrochanteric region of the femur was simulated in 12 rabbits. An oblong subtrochanteric defect of a standardized size (reducing the torsional strength of the bones approximately by 66%) was created and an intramedullary implant made of titanium or the FRC composite was inserted. The contralateral femur served as the intact control. At 12 weeks of healing, the femurs were harvested and analyzed by radiography, torsional testing, micro-CT imaging and hard tissue histology. The functional recovery was unremarkable in both groups, although the final analysis revealed two healed undisplaced peri-implant fractures in the group of FRC implants. FEM studies demonstrated differences in stress-shielding effects of the titanium and FRC implants, but the expected biological consequences did not become evident during the follow-up time of the animal study. Biomechanical testing of the retrieved femurs showed no significant differences between the groups. The torsional strength of the fixed bones had returned the level of contralateral intact femurs. Both implants showed ongrowth of intramedullary new bone. No adverse tissue reactions were observed. Based on these favorable results, a large-scale EU-project (NewBone, www.hb.se/ih/polymer/newbone) has been launched for development of orthopaedic FRC implants.

Bone attachment to glass-fibre-reinforced composite implant with porous surface.

A method has recently been developed for producing fibre-reinforced composites (FRC) with porous surfaces, intended for use as load-bearing orthopaedic implants. This study focuses on evaluation of the bone-bonding behaviour of FRC implants. Three types of cylindrical implants, i.e. FRC implants with a porous surface, solid polymethyl methacrylate (PMMA) implants and titanium (Ti) implants, were inserted in a transverse direction into the intercondular trabeculous bone area of distal femurs and proximal tibias of New Zealand White rabbits. Animals were sacrificed at 3, 6 and 12 weeks post operation, and push-out tests (n=5-6 per implant type per time point) were then carried out. At 12 weeks the shear force at the porous FRC-bone interface was significantly higher (283.3+/-55.3N) than the shear force at interfaces of solid PMMA/bone (14.4+/-11.0 N; p<0.001) and Ti/bone (130.6+/-22.2N; p=0.001). Histological observation revealed new bone growth into the porous surface structure of FRC implants. Solid PMMA and Ti implants were encapsulated mostly with fibrous connective tissue. Finite element analysis (FEA) revealed that porous FRC implants had mechanical properties which could be tailored to smooth the shear stress distribution at the bone-implant interface and reduce the stress-shielding effect.