Intravitreal Injection of Anti-VEGF Antibody Induces Glomerular Endothelial Cells Injury.

INTRODUCTION: Antiangiogenic agents that inhibit vascular endothelial growth factor have emerged as important tools in cancer therapy and ocular diseases. Their systemic use can induce renal limited microangiopathy. Local use of anti-VEGF agent is supposed to be safe. We report here a unique case of early endothelial cells injury induced by intravitreal injection of bevacizumab. CASE PRESENTATION: A 72-year-old man was addressed for acute kidney injury with proteinuria. He was under treatment with intravitreal injections of bevacizumab for glaucoma. Kidney biopsy was performed and electron microscopy showed signs of early stages of glomerular microangiopathy. Bevacizumab was discontinued resulting in the improvement of renal function and albuminuria. DISCUSSION: Bevacizumab, a humanized monoclonal antibody to VEGF is an approved therapy for metastatic cancer. Systemic adverse events including thrombotic microangiopathy have been mainly reported after its systemic injection. Podocytes produce VEGF that interacts with endothelial cells VEGF receptor-2 maintaining glomerular basement membrane integrity. Bevacizumab induce the detachment of endothelial cells from glomerular basement membrane leading to the proteinuria and renal function decline. Intravitreal bevacizumab is generally supposed to be safe. However, glomerular injury with microangiopathy features, even after intravitreal injection is possible. CONCLUSION: We report the electron microscopy evidence that intravitreal injection of anti-VEGF induces glomerular endothelial cells injury. Nephrologists and ophthalmologists should be aware of this complication.

Interplay of water and reactive elements in oxidation of alumina-forming alloys.

High-temperature alloys are crucial to many important technologies that underpin our civilization. All these materials rely on forming an external oxide layer (scale) for corrosion protection. Despite decades of research on oxide scale growth, many open questions remain, including the crucial role of the so-called reactive elements and water. Here, we reveal the hitherto unknown interplay between reactive elements and water during alumina scale growth, causing a metastable 'messy' nano-structured alumina layer to form. We propose that reactive-element-decorated, hydroxylated interfaces between alumina nanograins enable water to access an inner cathode in the bottom of the scale, at odds with the established scale growth scenario. As evidence, hydride-nanodomains and reactive element/hydrogen (deuterium) co-variation are observed in the alumina scale. The defect-rich alumina subsequently recrystallizes to form a protective scale. First-principles modelling is also performed to validate the RE effect. Our findings open up promising avenues in oxidation research and suggest ways to improve alloy properties.

Distribution of Silica-Coated Silver/Gold Nanostars in Soft- and Hardwood Applying SERS-Based Imaging.

Nanoparticles (NPs) in aqueous suspension have just begun to be exploited for the preservative treatment of wood. However, at present, there is very little information available on the distribution of NPs in wood after impregnation, due to associated analytical challenges. In this study, we present the detection of model NPs in softwood and hardwood by surface-enhanced Raman spectroscopy (SERS). SERS is a highly sensitive analytical method requiring no fluorescent labeling. The NP distribution after impregnation is evaluated with one representative species of the two wood types. To show the feasibility of the method, we prepared SERS-active Au/Ag nanostars coated with silica to act as a model NP system. We show herein that NPs can be imaged in very low quantities in both wood types without any matrix interactions. The presence of the NPs in the wood was confirmed by scanning electron microscopy (SEM) imaging and energy dispersive X-ray analysis (EDX). The fast detection of NPs in a complex matrix, without complicated sample preparation, marks a huge step forward in the development and application of nanotechnology for wood preservation and the quest to optimize the properties of one of the world's most important raw materials.

Severe right heart failure in a patient with chronic obstructive lung disease: a diagnostic challenge.

A 55-year-old male was admitted for evaluation of severe dyspnoea and hypoxaemia. Physical examination upon admission showed elevated jugular venous pressure and an accentuated second heart sound. Chest radiograph showed cardiomegaly with increased bibasilar markings. Arterial blood gas analysis while breathing room air showed marked hypoxaemia. High resolution computed tomography angiography of the chest showed modestly enlarged mediastinal lymph nodes with discrete diffuse ground-glass attenuation especially at the lower lung zones. Positron emission tomography using 18F labelled 2-deoxy-D-glucose (FDG) demonstrated the mediastinal lymph nodes were FDG-avid. Transthoracic echocardiography showed dilated hypokinetic right heart chambers with bulging of the interventricular septum to the left, compatible with acute cor-pulmonale. From the tricuspid regurgitation jet measurement a systolic pulmonary artery pressure (PAP) of 48 mmHg was estimated. Patent foramen ovale was suspected on bubble test. Right heart catheterisation confirmed pulmonary arterial hypertension: mPAP 47 mmHg, pulmonary artery occlusion pressure 5 mmHg, cardiac index 1.1 L/min/m2, pulmonary vascular resistance (PVR) 959 dyne.sec.cm(-5). Pulmonary function tests showed a marked diffusing capacity for carbon monoxide (DLCO) decrease of 32% predicted but no obstructive lung deficit. Before an open lung biopsy could be scheduled the patient developed acute cardiogenic shock. At autopsy pulmonary veno-occlusive disease with marked pulmonary hypertension was diagnosed.

Deep venous thrombosis and pulmonary embolism caused by an intravascular synovial sarcoma of the common femoral vein.

Malignant tumors arising in deep veins of the lower extremities are very uncommon. To our best knowledge, this is the seventh case of a primary venous intravascular synovial sarcoma (SS) reported in literature. A 32-year-old woman was admitted with a second episode of deep venous thrombosis of the right lower limb and pulmonary embolism. Physical and radiological examinations showed besides the thrombosis a tumor arising from the right common femoral vein involving the bifurcation of the common femoral artery. At surgery, en block resection of the tumor including the deep femoral vein and arterial bifurcation was done with an arterial reconstruction using a synthetic graft. Histopathological examination revealed an intravascular SS of the common femoral vein. The mainstay of curative therapy is complete surgical resection of all tumor manifestations with negative histological margins.

An incident case of primary acquired pulmonary alveolar proteinosis.

This is a case of a primary acquired pulmonary alveolar proteinosis (PAP) in an asymptomatic patient, on the waiting list for kidney transplantation, confirmed on lung biopsy and by identifying anti-GM-CSF antibodies.

Pre-terminal renal insufficiency in a patient with enteric hyperoxaluria: effect of medical management on renal function.

Enteric hyperoxaluria causes tubular deposition calcium oxalate crystals and severe chronic interstitial nephritis. We describe a patient with pre-terminal renal failure due to oxalate nephropathy after ileal resection. Increased oral hydration, low oxalate diet, and oral calcium carbonate and potassium citrate supplements resulted in a significant improvement of renal function. During the three-year follow-up, urinary oxalate concentration was repeatedly reduced below the crystallization threshold and serum creatinine decreased from 4.5 to 1.7 mg/dL. This case illustrates the benefit of combining and optimizing dietary and medical management in enteric hyperoxaluria, even in patients with advanced chronic kidney disease.

Clinical activity of afatinib (BIBW 2992) in patients with lung adenocarcinoma with mutations in the kinase domain of HER2/neu.

Human epidermal growth factor receptor (HER)2/neu kinase domain mutations are found in approximately 1-4% of lung adenocarcinomas with a similar phenotype to tumors with epidermal growth factor receptor (EGFR) mutations. Afatinib is a potent irreversible ErbB family blocker. We determined the tumor genomic status of the EGFR and HER2 genes in non- or light smokers with lung adenocarcinoma in patients who were entered into an exploratory Phase II study with afatinib. Five patients with a non-smoking history and metastatic lung adenocarcinomas bearing mutations in the kinase domain of HER2 gene were identified, three of which were evaluable for response. Objective response was observed in all three patients, even after failure of other EGFR- and/or HER2-targeted treatments; the case histories of these patients are described in this report. These findings suggest that afatinib is a potential novel treatment option for this subgroup of patients, even when other EGFR and HER2 targeting treatments have failed.

Sepsis-related acute kidney injury: a protective effect of drotrecogin alpha (activated) treatment?

BACKGROUND: Drotrecogin alpha activated (DrotAA) is licensed for treatment of patients with severe sepsis and organ failure. Among the latter, acute kidney injury (AKI), defined as the persistence of oligo-anuria following adequate resuscitation, is one of the most apprehended. We conducted a prospective, observational, and controlled study to test the hypothesis that DrotAA beneficially affected the evolution and outcome of AKI, complicating acute sepsis-induced cardiopulmonary failure. METHODS: Forty-six patients were studied. Thirty subjects received standard treatment for sepsis without DrotAA. In the remaining 16 patients, DrotAA was added as a continuous infusion of 24 microg/kg/h for 96 h. RESULTS: Mean age, causes of sepsis, and severity/organ failure scores were comparable between patients treated with or without DrotAA. Mortality at 28 days was high and comparable between both treatment groups (56% vs. 69%, DrotAA vs. no DrotAA; P=0.5). When oligo-anuria was present at the start of the study, it persisted during treatment in all patients, with no significant difference between groups. Both treatment groups presented with baseline mean daily fractional excretion of sodium values >2% that remained high during the observation period, regardless of whether DrotAA was given or not. Kidney histology showed a preserved renal architecture with tubular necrosis in all specimens. Similar glomerular, tubulo-interstitial, and vascular alterations were present in both treatment groups. CONCLUSION: In this small cohort of patients with severe sepsis who received adjuvant DrotAA treatment, no effect on urine output, tubular function, or mortality could be demonstrated.

The deep belly of the temporalis muscle: an anatomical, histological and MRI study.

In order to achieve a better functional and clinical knowledge of a masticatory muscle called the sphenomandibularis that is suspected to be responsible for headaches by compressing the maxillary nerve, bilateral dissections of the infratemporal fossa were performed on ten human cadavers and completed by histological and radiological studies of the same areas. Both macroscopic and microscopic observations obviously showed that the so-called sphenomandibularis muscle corresponds to the deep portion of the temporalis muscle, since there is no epimysial septum between these two structures, which previously have been described as being completely independent from each other. In spite of the close topographic relationship between the deep belly of the temporalis and the lateral pterygoid muscle, as well as their similar innervation pattern, the sphenomandibularis in fact has to be considered functionally as an original but non-isolated positional fascicle of the temporalis muscle itself. Our observations, correlated with MR images, suggest indeed that the deep belly of the temporalis muscle is of functional importance in the masticatory movements, but is not involved by its neurovascular vicinity in the genesis of specific headaches. Its surgical release, however, should be discussed in the case of a temporal myoplasty.

Carbon dioxide transport and carbonic anhydrase in blood and muscle.

CO(2) produced within skeletal muscle has to leave the body finally via ventilation by the lung. To get there, CO(2) diffuses from the intracellular space into the convective transport medium blood with the two compartments, plasma and erythrocytes. Within the body, CO(2) is transported in three different forms: physically dissolved, as HCO(3)(-), or as carbamate. The relative contribution of these three forms to overall transport is changing along this elimination pathway. Thus the kinetics of the interchange have to be considered. Carbonic anhydrase accelerates the hydration/dehydration reaction between CO(2), HCO(3)(-), and H(+). In skeletal muscle, various isozymes of carbonic anhydrase are localized within erythrocytes but are also bound to the capillary wall, thus accessible to plasma; bound to the sarcolemma, thus producing catalytic activity within the interstitial space; and associated with the sarcoplasmic reticulum. In some fiber types, carbonic anhydrase is also present in the sarcoplasm. In exercising skeletal muscle, lactic acid contributes huge amounts of H(+) and by these affects the relative contribution of the three forms of CO(2). With a theoretical model, the complex interdependence of reactions and transport processes involved in CO(2) exchange was analyzed.

Effects of carbonic anhydrase inhibitors on oxygen consumption and lactate accumulation in skeletal muscle.

In isolated rat soleus and extensor digitorum longus (EDL) muscles, the effects of carbonic anhydrase inhibitors were studied on oxygen consumption as well as lactate release and accumulation after incubation in inhibitors lasting long enough to produce marked changes in contractile parameters and in the concentrations of energy-rich phosphates. The inhibitors used were chlorzolamide (10(-3) M) and NaCNO (10(-2) M). Compared with control muscles, muscles treated with either of the two inhibitors showed a decrease in force, and an increase in time-to-peak as well as in relaxation time. Lactate content and release in soleus and in EDL were increased by factors of 2-3 with both inhibitors. With both inhibitors, oxygen consumption in the red soleus increased by approximately 27%, whereas in EDL, no significant change could be observed. The increase in aerobic metabolic rate in the red soleus only might indicate that the isozyme CA III, which is present only in this type of muscle, is in some way involved in keeping the oxygen consumption low. The increase in anaerobic metabolic rate occurring in both muscles can possibly be explained by increases in Pi and ADP.

Contractile function of papillary muscles with carbonic anhydrase inhibitors.

Contractile parameters of directly stimulated rabbit papillary muscles were studied during incubation in baths containing the carbonic anhydrase inhibitors chlorzolamide (1*10(-3) M) or ethoxzolamide (1*10(-4) M). Both inhibitors caused an at least partly reversible decrease in isometric force as it has been observed in skeletal muscle, and--in contrast to the results in skeletal muscles--a decrease in time-to-peak and half-relaxation time. It is postulated that inhibition of the membrane-bound carbonic anhydrase of heart muscle might induce an intracellular acidosis and that this acidosis causes the observed effects on contractile parameters.

Carbonic anhydrase in skeletal and cardiac muscle from rabbit and rat.

We have studied the distribution of carbonic anhydrases (CA) in several skeletal muscles of the hindlimb of rabbits and rats and in cardiac muscle of the rabbit. To remove erythrocyte CA, hindlimbs and hearts were thoroughly perfused with dextran solution, and the effectiveness of the perfusion was in most cases assessed by determining the contamination of the muscles with radioisotopes that had been used to label the erythrocytes before the perfusion was started. We observed three forms of CA: (1) cytosolic (sulphonamide-resistant) CA III; (2) a cytosolic sulphonamide-sensitive CA, probably isoenzyme II; (3) a membrane-bound form that was extracted from the particulate fraction using Triton X-100. These CA isoforms were distributed as follows. (1) CA III is located in the cytoplasm of slow, oxidative skeletal muscles and is absent from or low in fast skeletal and cardiac muscle; this holds for rabbits and rats and is identical with the pattern previously described for several other species. (2) The cytosolic sulphonamide-sensitive CA is present in fast rabbit muscles and absent from slow muscles of this species. In contrast, all skeletal muscles of the rat studied here lack, or possess only very low, activity of this isoenzyme. (3) The membrane-bound form of CA is present in all rabbit muscles studied; its activity appears somewhat higher in fast than in slow skeletal muscles. (4) Cardiac muscle constitutes an exception among all striated muscles of the rabbit as it possesses no form of cytosolic CA but a high activity of the membrane-bound form.

Effects of carbonic anhydrase inhibitors on contraction, intracellular pH and energy-rich phosphates of rat skeletal muscle.

1. The effects of carbonic anhydrase inhibitors on contractile parameters, intracellular pH (pHi) and energy-rich phosphates were studied in isolated rat soleus and extensor digitorum longus (EDL) muscles. 2. The muscles were incubated either in Ringer solutions (95% O2/5% CO2 = control) or in solutions to which one of the inhibitors, 5 X 10(-4) M-chlorzolamide or 10(-2) M-NaCNO, had been added. Muscles were stimulated directly and contracted under isometric conditions. 3. Compared with control muscles, both inhibitor-treated muscles showed a significantly decreased tetanic force and an increased half-relaxation time of twitches and tetani. Chlorzolamide increased time-to-peak in both muscles. Cyanate decreased isometric twitch force in both muscles. 4. Both inhibitors decreased pHi in both muscles; chlorzolamide by 0.1 unit, cyanate by 0.4 unit in soleus and by 0.8 unit in EDL. 5. Chlorzolamide increased the concentrations of creatine and inorganic phosphate (Pi) in soleus (the effect of chlorzolamide was not studied in EDL). Cyanate caused these same changes in soleus as well as EDL and in addition decreased the concentrations of ATP and phosphocreatine in soleus and EDL. 6. In muscles acidified by either low external HCO3- (2 mM) or by elevated PCO2 (30% CO2 in the gas phase) in the bath, decreases in isometric force and increases in half-relaxation time of tetani were observed. In addition there were increases in muscle Pi. These effects were more pronounced with 30% CO2 than with 2 mM-HCO3-. 7. Neither acidifying solutions prolonged either half-relaxation time or time-to-peak of twitches. 8. We conclude that carbonic anhydrase inhibition exerts its effect (a) on isometric tension at least partly via an elevated Pi (perhaps in combination with lowered pHi); (b) on the half-relaxation time of tetani by means of lowered pHi and elevated concentration of Pi; (c) on relaxation and time-to-peak of twitches by some unknown mechanism, neither directly by a change in pHi nor in Pi.

Carbonic anhydrase inhibition affects contraction of directly stimulated rat soleus.

We have studied the contractile parameters of directly stimulated isolated rat soleus muscles incubated in media containing the carbonic anhydrase inhibitors chlorzolamide (5.10(-4)M) or cyanate (10(-2)M). Both inhibitors caused a decrease in isometric twitch and tetanic (5s) tensions and an increase in muscle relaxation time. It is speculated that among the three types of skeletal muscle carbonic anhydrase it may be the enzyme associated with the sarcoplasmatic reticulum whose inhibition caused the observed changes in contractile parameters.

Effects of dextran-bound inhibitors on carbonic anhydrase activity in isolated rat lungs.

Effects of macromolecular Prontosil-dextran inhibitors (PD) on carbonic anhydrase (CA) activity in isolated rat lungs were studied. Isolated lungs were perfused with Krebs-Ringer bicarbonate (KRB) solutions containing no inhibitor, PD 100,000 (mol wt 100,000), PD 5,000 (mol wt 5,000), or low-molecular-weight inhibitors (Prontosil or acetazolamide). The time course of effluent perfusate pH equilibration was measured in a stop-flow pH electrode apparatus. Pulmonary CO2 excretion (Vco2) was monitored by continuously recording expired CO2 concentration. The lungs were ventilated with room air and perfused at 37 degrees C with KRB prebubbled with 5% CO2- 20% O2- 75% N2. The results obtained show that both the low-molecular-weight inhibitors and PD's caused postcapillary pH disequilibria (delta pH) in effluent perfusate. However, only acetazolamide and Prontosil caused a reduction in Vco2. These results suggest that there is an intravascular CA, presumably associated with endothelial cell membranes, that is accessible to all inhibitors used and is responsible in part for equilibration of the CO2- HCO3- -H+ reactions in the perfusate but, under the conditions used, does not affect CO2 excretion; and there is an extravascular (possibly intracellular) CA that can be inhibited by low-molecular-weight inhibitors, is primarily responsible for enhanced CO2 transfer across the alveolar-capillary barrier (perhaps via facilitation of CO2 diffusion), and is in part responsible for pH equilibration.

Extracellular carbonic anhydrase of skeletal muscle associated with the sarcolemma.

We report here 1) the synthesis and properties of a new macromolecular carbonic anhydrase inhibitor, Prontosil-dextran, 2) its application to determine the localization of a previously described extracellular carbonic anhydrase in skeletal muscle, and 3) the application of a recently published histochemical technique using dansylsulfonamide to the same problem. Stable macromolecular inhibitors of molecular weights of 5,000, 100,000 and 1,000,000 were produced by covalently coupling the sulfonamide Prontosil to dextrans. Their inhibition constants towards bovine carbonic anhydrase II are 1-2 X 10(-7) M. The Prontosil-dextrans, PD 5,000, PD 100,000, and PD 1,000,000, were used in studies of the washout of H14CO3-) from the perfused rabbit hindlimb. This washout is slow due to the presence of an extracellular carbonic anhydrase and can be markedly accelerated by PD 5,000 but not by PD 100,000 and PD 1,000,000. Since PD 5,000 is accessible to the entire extracellular space and PD 100,000 and PD 1,000,000 are confined to the intravascular space, we conclude that the extracellular carbonic anhydrase of skeletal muscle is located in the interstitium. The histochemical studies show a strong staining of the sarcolemma of the muscle fibers with high oxidative capacity. It appears likely, therefore, that the extracellular carbonic anhydrase of skeletal muscle is associated with muscle plasma membranes with its active site directed toward the interstitial space.

Inhibition properties and inhibition kinetics of an extracellular carbonic anhydrase in perfused skeletal muscle.

We have investigated the properties of the carbonic anhydrase which is functionally available to CO2 and HCO3- in the capillary bed of skeletal muscle. We used essentially the indicator-dilution technique of Effros and Weissman (J. Appl. Physiol. 47, 1090-1098, 1979). Into hindlimbs of rabbits perfused with dextran solution we injected boli containing H14CO3- or 36Cl-, and 3H-dextran (MW 80 000) as an intravasal indicator, and observed the washout of these indicators by fractionated collection and analysis of the venous effluent. In agreement with previous studies we found that addition of 10(-4) M of the carbonic anhydrase inhibitor acetazolamide to the perfusate considerably speeds up the washout of 14C, reducing the extraction of 14C from 0.72 to 0.45. A half-maximal effect on 14C extraction was achieved with 1 x 10(-6) M acetazolamide (IC50). The carbonic anhydrase inhibitors methazolamide and benzolamide both yielded IC50 values of 10(-5) M. This pattern of inhibitory potency of the three sulfonamides is incompatible with their inhibitory effects on the two known cytosolic isoenzymes of skeletal muscle, CAII and CAIII. While cells take up acetazolamide and benzolamide extremely slowly, with half-times of several minutes to hours, the effect of both sulfonamides on 14C washout occurred very rapidly: less than 1 min, probably not more than a few seconds, were necessary to achieve inhibitory effects. We conclude that (1) a tissue carbonic anhydrase converts the injected H14CO3- quickly into 14CO2 which then diffuses into the intracellular space thus causing a washout of 14C that is much slower than that of the intravasal indicator or that of 36Cl-, (2) this carbonic anhydrase is not intra- but extracellular and presumably membrane-bound, and (3) its properties suggest that it is distinct from the well-known cytosolic carbonic anhydrases and represents a different isoenzyme.