Canal wall down mastoidectomy with obliteration versus canal wall up mastoidectomy in primary cholesteatoma surgery.

OBJECTIVE: This study sought to compare disease recidivism rates between canal wall up mastoidectomy and a canal wall down with obliteration technique. METHODS: Patients undergoing primary cholesteatoma surgery at our institution over a five-year period (2013-2017) using the aforementioned techniques were eligible for inclusion in the study. Rates of discharge and disease recidivism were analysed using chi-square statistics. RESULTS: A total of 104 ears (98 patients) were included. The mean follow-up period was 30 months (range, 12-52 months). A canal wall down with mastoid obliteration technique was performed in 55 cases and a canal wall up approach was performed in 49 cases. Disease recidivism rates were 7.3 per cent and 16.3 per cent in the canal wall down with mastoid obliteration and canal wall up groups respectively (p = 0.02), whilst discharge rates were similar (7.3 per cent and 10.2 per cent respectively). CONCLUSION: Our direct comparative data suggest that canal wall down mastoidectomy with obliteration is superior to a canal wall up technique in primary cholesteatoma surgery, providing a lower recidivism rate combined with a low post-operative ear discharge rate.

Effect of pulsed electromagnetic fields on endoplasmic reticulum stress.

The maintenance of protein homeostasis in the endoplasmic reticulum (ER) is crucial in cell life. Disruption of proteostasis results in ER stress that activates the unfolded protein response (UPR); a signalling network assigned to manage the accumulated misfolded or unfolded proteins. Prolonged or unresolved ER stress leads to apoptotic cell death that can be the basis of many serious diseases. Our aim was to study the effect of pulsed electromagnetic fields (PEMF), an alternative, non-invasive therapeutic method on ER stressed cell lines. First, the effect of PEMF treatment on the expression of ER stress markers was tested in three different cell lines. PEMF had no remarkable effect on ER stress protein levels in human embryonic kidney (HEK293T) and human liver carcinoma (HepG2) cell lines. However, the expression of BiP, Grp94 and CHOP were increased in HeLa cells upon PEMF exposure. Therefore, HepG2 cell line was selected for further experiments. Cells were stressed by tunicamycin and exposed to PEMF. Grp94, PDI, CHOP and PARP expression as markers of stress were monitored by Western blot and cell viability was also investigated. Tunicamycin treatment, as expected, increased the expression of Grp94, PDI, CHOP and inactivated PARP. Analysis of protein expression showed that PEMF was able to decrease the elevated level of ER chaperons Grp94, PDI and the apoptosis marker CHOP. The truncated, inactive form of PARP was also decreased. Accordingly, cell viability was also improved by PEMF exposure. These results indicate that PEMF is able to moderate ER stress induced by tunicamycin in HepG2 cells. However, our results clearly draw attention to that different cell lines may vary in the response to PEMF treatment.

Characterisation of patients with endoscopy-negative, computer tomography-negative midfacial segment pain using the sino-nasal outcome test.

BACKGROUND: The purpose of this study was to qualitatively characterise patients with midfacial segment pain (MSP) using the Sino-Nasal Outcome Test (SNOT). The data will provide a detailed overview of the physical and psychological impact on patients'well-being, and how it compares with the normal, healthy population. METHODS: Suitable patients were prospectively identified from the Multi-disciplinary Facial Pain Clinic at the Royal Liverpool University Hospital, based on the diagnostic criteria for MSP. The pre-treatment SNOT-22 of these patients were also compared to patients with chronic rhinosinusitis and normal healthy volunteers. RESULTS: Twenty-nine consecutive patients with a diagnosis of MSP were identified, and compared with 30 CRS patients and 34 healthy volunteers. The average SNOT-22 scores of MSP and CRS patients were higher than normal healthy volunteers. Patients with CRS had the highest rhinological subscale SNOT scores compared to normal healthy volunteers and MSP. Conversely, the reported ear and facial symptoms of MSP patients were most unfavourable. A similar trend was observed in reported sleep function where MSP patients recorded higher subscale scores than the other two cohorts. The subscale mean score for psychological function of MSP patients was not significant when compared to the mean score of patients diagnosed with CRS. CONCLUSION: MSP has an adverse impact on both physical and psychological well-being. The subtle differences in the SNOT subscores between MSP and CRS have provided greater insight into the character and disease impact of MSP. We propose that the SNOT may be suitably utilised in MSP to document disease severity and measure response to treatment.

Vitamin C: update on physiology and pharmacology.

Although ascorbic acid is an important water-soluble antioxidant and enzyme cofactor in plants and animals, humans and some other species do not synthesize ascorbate due to the lack of the enzyme catalyzing the final step of the biosynthetic pathway, and for them it has become a vitamin. This review focuses on the role of ascorbate in various hydroxylation reactions and in the redox homeostasis of subcellular compartments including mitochondria and endoplasmic reticulum. Recently discovered functions of ascorbate in nucleic acid and histone dealkylation and proteoglycan deglycanation are also summarized. These new findings might delineate a role for ascorbate in the modulation of both pro- and anti-carcinogenic mechanisms. Recent advances and perspectives in therapeutic applications are also reviewed. On the basis of new and earlier observations, the advantages of the lost ability to synthesize ascorbate are pondered. The increasing knowledge of the functions of ascorbate and of its molecular sites of action can mechanistically substantiate a place for ascorbate in the treatment of various diseases.

Endoplasmic reticulum stress underlying the pro-apoptotic effect of epigallocatechin gallate in mouse hepatoma cells.

It has been recently reported that tea flavanols, including epigallocatechin gallate (EGCG), efficiently inhibit glucosidase II in liver microsomes. Since glucosidase II plays a central role in glycoprotein processing and quality control in the endoplasmic reticulum we investigated the possible contribution of endoplasmic reticulum stress and unfolded protein response (UPR) to the pro-apoptotic activity of EGCG in mouse hepatoma cells. The enzyme activity measurements using 4-methylumbelliferyl-alpha-d-glucopyranoside substrate confirmed the inhibition of glucosidase II in intact and alamethicin-permeabilized cells. EGCG treatment caused a progressive elevation of apoptotic activity as assessed by annexin staining. The induction of CHOP/GADD153, the cleavage of procaspase-12 and the increasing phosphorylation of eIF2alpha were revealed in these cells by Western blot analysis while the induction of endoplasmic reticulum chaperones and foldases was not observed. Time- and concentration-dependent depletion of the endoplasmic reticulum calcium stores was also demonstrated in the EGCG-treated cells by single-cell fluorescent detection. The massive alterations in the endoplasmic reticulum morphology revealed by fluorescent microscopy further supported the development of UPR. Collectively, our results indicate that EGCG interferes with protein processing in the endoplasmic reticulum presumably due to inhibition of glucosidase II and that the stress induces an incomplete unfolded protein response with dominantly pro-apoptotic components.

Different expression and distribution of 11beta-hydroxysteroid dehydrogenase type 1 in obese and lean animal models of type 2 diabetes.

11beta-hydroxysteroid dehydrogenase type 1 (11betaHSD1) is a NADPH dependent oxidoreductase of the endoplasmic reticulum lumen which converts cortisone to cortisol and plays a role in the pathogenesis of metabolic syndrome and type 2 diabetes. The aim of our study was to investigate the correlation between the expression/activity of 11betaHSDI and obesity. Liver and adipose tissue microsomes of an obese (Zucker) and a non-obese (Goto-Kakizaki) type 2 diabetes model rat strains were used. 11betaHSDI expression was detected at mRNA, protein and activity level. The activity of 11betaHSD1 was increased in the adipose tissue and decreased in the liver of the obese Zucker rat, while its mRNA levels were significantly different only in the adipose tissue. In diabetic Goto-Kakizaki rat both the expression and the activity of 11betaHSD1 were elevated in liver, but not in adipose tissue. These results suggest that the prereceptorial glucocorticoid activation is different in the liver and adipose tissue of the two diabetes models. This phenomenon might be responsible for the obese and lean phenotypes in type 2 diabetes.

Dehydroascorbate and glucose are taken up into Arabidopsis thaliana cell cultures by two distinct mechanisms.

The possible involvement of glucose (Glc) carriers in the uptake of vitamin C in plant cells is still a matter of debate. For the first time, it was shown here that plant cells exclusively take up the oxidised dehydroascorbate (DHA) form. DHA uptake is not affected by 6-bromo-6-deoxy-ascorbate, an ascorbate (ASC) analogue, specifically demonstrating ASC uptake in animal cells. There is no competition between Glc and DHA uptake. Moreover, DHA and Glc carriers respond in the opposite manner to different inhibitors (cytochalasin B, phloretin and genistein). In conclusion, the plant plasma membrane DHA carrier is distinct from the plant Glc transporters.

Foreign bodies in the ear: a simple technique for removal analysed in vitro.

OBJECTIVE: Foreign bodies in the ear are mainly encountered in children. This can often pose a problem especially in an accident and emergency department where a microscope or expert help is not routinely available. This paper presents a simple, safe, and effective way of ear syringing. The ease and simplicity of the procedure along with the equipment are described. METHOD AND RESULT: The equipment consists of a "disposable" sterile kit, consisting of a 20 ml syringe, saline at body temperature and 14 or 16 gauge cannula (without the needle). An in vitro experiment was conducted to calculate the pressure generated by the water jet on the eardrum. The pressure was well below the pressure required to burst a tympanic membrane, and hence this technique is safe to use. CONCLUSIONS: Ear syringing is an effective and easy way of removing most foreign bodies. A detailed history and an otoscopic examination must precede the procedure. The novel method of syringing described in this paper with the usual safeguards could be a useful adjunct in the management of this common condition.

The regulation of the induction of vascular endothelial growth factor at the onset of diabetes in spontaneously diabetic rats.

Early induction of VEGF was studied in liver, kidney and lung of spontaneously diabetic rats. Western blot analysis, northern hybridization were applied to show the expression of VEGF in different organs. Radiolabelled hypoxia responsive element (HRE) and cAMP responsive element (CRE) oligonucleotides were assayed by electrophoretic mobility shift (EMSA) or supershift using anti ARNT and anti CREB-1 monoclonal antibodies. An increase in VEGF expression at the level of protein and mRNA was demonstrated at the beginning of the disease. EMSAs showed: a.) a binding of HIF-1 to HRE and/or CRE, b.) in the same time the binding of CREB- I was detected to both HRE and/or CRE sequences in the liver, kidney and lung of diabetic animals. Based on these in vivo observations it is supposed that HRE and CRE through the interaction between HIF-1 and CREB-1 are equally involved in the regulation of VEGF expression at the onset of diabetes.

Ryanodine receptor channel-dependent glutathione transport in the sarcoplasmic reticulum of skeletal muscle.

We found that glutathione transport across endo/sarcoplasmic reticulum membranes correlates with the abundance of ryanodine receptor type 1 (RyR1). The transport was the fastest in muscle terminal cisternae, fast in muscle microsomes and slow in liver, heart, and brain microsomes. Glutathione influx could be inhibited by RyR1 blockers and the inhibitory effect was counteracted by RyR1 agonists. The effect of blockers was specific to glutathione, as the transport of other small molecules was not hindered. Therefore, the glutathione transport activity seems to be associated with RyR1 in sarcoplasmic reticulum.

Glucose 6-phosphate transport in fibroblast microsomes from glycogen storage disease type 1b patients: evidence for multiple glucose 6-phosphate transport systems.

In liver endoplasmic reticulum the intralumenal glucose-6-phosphatase activity requires the operation of a glucose 6-phosphate transporter (G6PT1). Mutations in the gene encoding G6PT1 cause glycogen storage disease type 1b, which is characterized by a loss of glucose-6-phosphatase activity and impaired glucose homoeostasis. We describe a novel glucose 6-phosphate (G6P) transport activity in microsomes from human fibroblasts and HeLa cells. This transport activity is unrelated to G6PT1 since: (i) it was similar in microsomes of skin fibroblasts from glycogen storage disease type 1b patients homozygous for mutations of the G6PT1 gene, and in microsomes from human control subjects; (ii) it was insensitive to the G6PT1 inhibitor chlorogenic acid; and (iii) it was equally active towards G6P and glucose 1-phosphate, whereas G6PT1 is highly selective for G6P. Taken together, our results provide evidence for the presence of multiple transporters for G6P (and other hexose phosphoesters) in the endoplasmic reticulum.

The hepatic glycogenoreticular system.

One of the major liver functions is the ability of hepatocytes to store glucose in the form of glycogen for various purposes. Beside glucose production and secretion, the synthesis of glucuronides and ascorbate has been reported to be dependent on the extent of the glycogen stores and on the rate of glycogenolysis in the liver. It is common that the final steps of these pathways are catalysed by intraluminally orientated enzymes of the endoplasmic reticulum, which are supported by transporters for the permeation of substrates and products. On the basis of the close morphological and functional proximity of glycogen, glycogen-dependent pathways and the (smooth) endoplasmic reticulum we propose to use the term glycogenoreticular system for the description of this export-orientated hepatocyte-specific metabolic unit.

Endonasal laser dacryocystorhinostomy and external dacryocystorhinostomy outcome profile in a general ophthalmic service unit: a comparative retrospective study.

PURPOSE: To evaluate the outcome profile of endonasal laser dacryocystorhinostomy (ENL-DCR) in comparison with external dacryocystorhinostomy (ENL-DCR) carried out as part of general ophthalmic service within the same center. METHODS: Patients who have undergone external or endonasal laser DCR in the authors institute with a minimum follow-up of 9 months and at least 3 months after removal of the tubes were invited to participate in this research. We used a questionnaire and a systematic clinical examination for detecting lacrimal passage patency and function. Patients were classified into categories: complete anatomical and physiological success; anatomical success with partial relief of symptoms; anatomical success with no relief of symptoms; anatomical failure. The endoscopic view of the ostium vertical location has been classified into four levels. RESULTS: One hundred and ten external-DCR and 53 Endonasal-DCR procedures were evaluated. Free communication (anatomical success) was achieved in 82% undergoing Ext-DCR and in 58% undergoing ENL-DCR. A significant number of patients continued to have symptoms in spite of a patent fistula (54% for Ext-DCR and 39% for ENL-DCR). The site of the opening of the internal ostium was significantly related to the persistence of symptoms in spite of free communication (P < 0.001, chi-square test). CONCLUSION: In this series of patients undergoing DCR in a general ophthalmic unit, the standard Ext-DCR technique has a higher anatomical success rate than the endoscopic laser DCR but not necessarily with equivalent rate of relief of symptoms. An inferiorly placed ostium is more likely to result in complete relief of symptoms.

Role of vitamin E in ascorbate-dependent protein thiol oxidation in rat liver endoplasmic reticulum.

Addition of ascorbate or its generation from gulonolactone causes the oxidation of protein thiols and a simultaneous dehydroascorbate formation in rat liver microsomes. The participation of vitamin E in the phenomenon was studied. We measured ascorbate and protein thiol oxidation and lipid peroxidation in vitamin E deficient liver microsomes. Vitamin E deficiency partly uncoupled the two processes: ascorbate oxidation increased, while protein thiol oxidation decreased. These changes were accompanied with an accelerated lipid peroxidation in the vitamin E-deficient microsomes, which indicates the accumulation of reactive oxygen species. All these effects were reduced by the in vitro addition of vitamin E to the deficient microsomes, supporting its direct role in the process. The results demonstrate that vitamin E is a component of the protein thiol oxidizing machinery in the hepatic endoplasmic reticulum transferring electrons from the thiol groups towards oxygen.

Protein-disulfide isomerase- and protein thiol-dependent dehydroascorbate reduction and ascorbate accumulation in the lumen of the endoplasmic reticulum.

The transport and intraluminal reduction of dehydroascorbate was investigated in microsomal vesicles from various tissues. The highest rates of transport and intraluminal isotope accumulation (using radiolabeled compound and a rapid filtration technique) were found in hepatic microsomes. These microsomes contain the highest amount of protein-disulfide isomerase, which is known to have a dehydroascorbate reductase activity. The steady-state level of intraluminal isotope accumulation was more than 2-fold higher in hepatic microsomes prepared from spontaneously diabetic BioBreeding/Worcester rats and was very low in fetal hepatic microsomes although the initial rate of transport was not changed. In these microsomes, the amount of protein-disulfide isomerase was similar, but the availability of protein thiols was different and correlated with dehydroascorbate uptake. The increased isotope accumulation was accompanied by a higher rate of dehydroascorbate reduction and increased protein thiol oxidation in microsomes from diabetic animals. The results suggest that both the activity of protein-disulfide isomerase and the availability of protein thiols as reducing equivalents can play a crucial role in the accumulation of ascorbate in the lumen of the endoplasmic reticulum. These findings also support the fact that dehydroascorbate can act as an oxidant in the protein-disulfide isomerase-catalyzed protein disulfide formation.

Transcriptional induction of bilirubin UDP-glucuronosyltransrase by ethanol in rat liver.

Several drug-metabolizing enzymes including bilirubin UDP-glucuronosyltransferase (UGT1A1) are influenced by long-term ethanol consumption. In the present study, the activity and expression of UGT1A1 were investigated in livers of ethanol-treated rats. Animals were treated daily for 15 days with ethanol or isocaloric amount of glucose solution by gastric intubation. Microsomes and total RNA were prepared from the liver of rats and analyzed by Western blot and Northern hybridization using UGT1A1 specific antibody and cDNA probe. Microsomal bilirubin UGT activity was also measured. The elevation of UGT1A1 mRNA was observed in the liver of ethanol consumer animals with the simultaneous increase in microsomal UGT1A1 protein leading to stimulated bilirubin glucuronidation both in vivo and in microsomal vesicles. These results arise the possibility of the transcriptional induction and/or the mRNA stabilization by ethanol consumption, which can be caused by ethanol itself or the metabolic changes due to the treatment.

Pars tensa retraction pockets in children: treatment by excision and ventilation tube insertion.

Tympanic membrane retraction pockets involving the pars tensa are not uncommon in clinical practice. Recurrent infections, ossicular erosion and cholesteatoma are the recognized sequelae. The management options include surveillance, medical treatment and surgery. The surgical procedures range from grommet insertion to extensive tympanoplasty procedures. We report our experience with simple excision and grommet insertion, performed in 31 ears in 26 patients as day cases. The follow-up ranged from 8 to 34 months with a mean of 16 months. The procedure was successful in 23 ears (success rate of 74%). Recurrence of retraction occurred in seven ears and in one ear there was a persistent perforation. Age, previous grommet insertion and severity of retraction did not have a statistically significant influence on the final outcome. We conclude that excision and grommet insertion is a simple, safe and efficient procedure for the management of tympanic membrane retraction pockets and can be considered in preference to extensive tympanoplasty.

Glucose-6-phosphatase in the insulin secreting cell line INS-1.

The glucose-6-phosphatase system of the glucose sensitive insulin secreting rat insulinoma cells (INS-1) was investigated. INS-1 cells contain easily detectable levels of glucose-6-phosphatase enzyme protein (assessed by Western blotting) and have a very significant enzymatic activity. The features of the enzyme (Km and Vmax values, sensitivity to acidic pH, partial latency, and double immunoreactive band) are similar to those of the hepatic form. On the other hand, hardly detectable levels of glucose-6-phosphatase activity and protein were present in the parent glucose insensitive RINm5F cell line. The mRNA of the glucose-6-phosphate transporter was also more abundant in the INS-1 cells. The results support the view that the glucose-6-phosphatase system of the beta-cell is associated with the regulation of insulin secretion.