IgG4-related disease (IgG4-RD) is an immune-mediated fibroinflammatory condition that can affect multiple organs and lead to tumefactive, tissue-destructive lesions. Reports have described inflammatory aortitis and periaortitis, the latter in the setting of retroperitoneal fibrosis (RPF), but have not distinguished adequately between these 2 manifestations. The frequency, radiologic features, and response of vascular complications to B cell depletion remain poorly defined. We describe the clinical features, radiology findings, and treatment response in a cohort of 36 patients with IgG4-RD affecting large blood vessels.Clinical records of all patients diagnosed with IgG4-RD in our center were reviewed. All radiologic studies were reviewed. We distinguished between primary large blood vessel inflammation and secondary vascular involvement. Primary involvement was defined as inflammation in the blood vessel wall as a principal focus of disease. Secondary vascular involvement was defined as disease caused by the effects of adjacent inflammation on the blood vessel wall.Of the 160 IgG4-RD patients in this cohort, 36 (22.5%) had large-vessel involvement. The mean age at disease onset of the patients with large-vessel IgG4-RD was 54.6 years. Twenty-eight patients (78%) were male and 8 (22%) were female. Thirteen patients (36%) had primary IgG4-related vasculitis and aortitis with aneurysm formation comprised the most common manifestation. This affected 5.6% of the entire IgG4-RD cohort and was observed in the thoracic aorta in 8 patients, the abdominal aorta in 4, and both the thoracic and abdominal aorta in 3. Three of these aneurysms were complicated by aortic dissection or contained perforation. Periaortitis secondary to RPF accounted for 27 of 29 patients (93%) of secondary vascular involvement by IgG4-RD. Only 5 patients demonstrated evidence of both primary and secondary blood vessel involvement. Of those treated with rituximab, a majority responded positively.IgG4-RD is a distinctive, unique, and treatable cause of large-vessel vasculitis. It can also involve blood vessels secondary to perivascular tumefactive lesions. The most common manifestation of IgG4-related vasculitis is aortitis with aneurysm formation. The most common secondary vascular manifestation is periaortitis with relative sparing of the aortic wall. Both primary vasculitis and secondary vascular involvement respond well to B cell depletion therapy. Immunohistochemistry studies reveal a preponderance of IgG4-positive plasma cells. Recognition of this condition as a distinct entity stemmed from the investigation of autoimmune pancreatitis, which was ultimately found to be part of a larger systemic process. Over the last decade, IgG4-RD has been reported to involve nearly every organ system.IgG4-related disease (IgG4-RD) is an immune-mediated fibroinflammatory condition that can affect multiple organs and lead to tumefactive, tissue-destructive lesions. It was not until 2008, however, that large-vessel involvement by IgG4-RD was recognized in the form of “inflammatory abdominal aortic aneurysm.” A variety of other macrovascular manifestations of IgG4-RD have been described since then, including coronary periarteritis, inflammatory thoracic aortic aneurysms, and periaortitis/arteritis in the setting of retroperitoneal fibrosis (RPF).,9 Such vascular manifestations can be associated with morbidity and mortality in the form of aortic dissection and sudden cardiac death. IgG4-RD was recognized as a potential cause of large-vessel vasculitis in the 2012 Chapel Hill Consensus Conference on Nomenclature of Vasculitides but remains a poorly understood entity .Microscopic vascular findings of IgG4-RD—namely, obliterative phlebitis, and obliterative arteritis—have been recognized since 2003.Review of the literature suggests that 2 forms of large-vessel disease may exist in IgG4-RD: a primary vasculitis and a secondary form of vascular involvement characterized by periaortic or periarterial involvement. However, most literature on IgG4-related large-vessel involvement to date has not distinguished clearly between these 2 forms. We studied a cohort of 160 IgG4-RD patients whose disease features have been characterized carefully. Among these patients, 36 (22.5%) were recognized as having features of vascular involvement.22.1 In contrast, a probable histologic diagnosis requires at least one of these histopathology features in conjunction with supportive clinical findings. The presence of nonobliterative phlebitis and mild to moderate degrees of tissue eosinophilia, both considered to be minor pathologic features, can strengthen the likelihood of IgG4-RD.This study was approved by the institutional review board of the Massachusetts General Hospital. All patients provided written consent to participate. The diagnosis of IgG4-RD hinges on the correlation of clinical findings with both specific histopathologic features and an increased number of IgG4+ plasma cells (or IgG4+/IgG+ ratio) in the affected tissue. The major histopathologic features include a dense lymphoplasmacytic infiltrate, obliterative phlebitis, and fibrosis that has a “storiform”  pattern in focal or diffuse areas. The presence of 2 of these features is highly suggestive of an IgG4-RD diagnosis.In addition, a cutoff IgG4+ plasma cells per high-power field (hpf) ranging from 10 to 200 cells/hpf (depending on the biopsied organ) and an IgG4+/IgG+ ratio of at least 0.40 was considered essential to the histopathologic diagnosis. Because elevated IgG4+ plasma cells have been documented with other etiologies of aortitis, a density of 50 cells/400× hpf and an IgG4+/IgG+ ratio of at least 0.50 was required for cases relying on aortic tissue to establish the diagnosis. All of the above histologic definitions are in accordance with international consensus criteria on the pathology of IgG4-RD. All serum IgG4 concentration measurements were performed with a serial dilution protocol to prevent the prozone phenomenon.Flow cytometry, performed on all samples at the time of patients’ presentations with active, untreated disease, was used to measure the absolute plasmablast count per milliliter by gating peripheral blood for CD19lowCD20−CD38+CD27+. Methods for collecting plasmablasts have been described.Inflammatory vascular involvement was defined by objective radiologic or pathologic findings in all patients except 2, who were diagnosed on clinical grounds. Patient #1 was a 35-year-old man with a carotid occlusion and biopsy-proven IgG4-related RPF. Patient #4 involved an aortic dissection in a 32-year-old man with no atherosclerotic risk factors and a negative genetic evaluation for Ehlers–Danlos syndrome. He had a moderate serum IgG4 concentration elevation and the single slide available from the aortic surgery demonstrated a lymphoplasmacytic infiltrate.2.2Electronic records of all patients were reviewed. Data pertaining to demographics, comorbidities, clinical manifestations, organ involvement, laboratory studies, pathology, and treatment were included. Relevant comorbid conditions such as hypertension, hyperlipidemia, diabetes mellitus, chronic kidney disease, and arterial disease in the coronary, carotid, or extremity vessels were recorded. Laboratory data before and after rituximab administration (at 3 and 6 months) were also collected.2.3All available vascular radiology data were reviewed and interpreted independently by 2 vascular radiologists. The radiology studies included computed tomography (CT) angiography, magnetic resonance (MR) angiography, and positron emission tomography (PET).18fluorodeoxyglucose (FDG) avidity within the blood vessel wall on PET. For the latter 2 features, the intensity of these findings were required to exceed that of typical atherosclerotic disease.Primary IgG4-related vasculitis was defined by the presence of 1 or more of the following: vessel wall thickening; vessel wall enhancement on contrast study; or minimal vascular wall findings. The main radiologic findings were: enhancing soft tissue adjacent to or encasing the vessel; minimal vascular wall thickening or enhancement; 18FDG avidity in the perivascular region; and minimal luminal narrowing.Secondary IgG4-related vascular involvement was defined by abnormal perivascular soft tissue with Patients with coronary artery involvement were defined by either abnormal FDG uptake on PET imaging or large coronary artery aneurysms with circumferential mural thickening without concurrent evidence of atherosclerotic plaque on coronary CT angiography. Carotid arteritis was defined clinically as explained above in Patient #1, histopathologically in another, and radiographically in a 3rd with both contrast enhancement and FDG avidity of the vessel wall. Iliac phlebitis was defined by the presence of abnormal T2 hyperintensity on MR imaging along the iliac veins and associated abnormal FDG uptake on PET imaging. The presence of aortic or common iliac arterial aneurysms as well as bilateral hydroureteronephrosis, ureteral displacement, and lymphadenopathy were also evaluated. Thoracic aortic aneurysm was defined as a double oblique short-axis diameter of greater than 3.8 cm in the ascending thoracic aorta and greater than 3.0 cm in the descending thoracic aorta. Common iliac artery aneurysm was defined as a double oblique short-axis diameter of greater than 1.5 cm. Radiologic improvement following treatment was determined by either a decrease in thickening of the blood vessel wall, perivascular soft tissue thickening, contrast enhancement, or FDG avidity.Abdominal aortic aneurysm was defined as a double oblique short-axis diameter of greater than 3.0 cm.2.4Aortitis was defined as inflammation of the adventitia of the aorta with at least focal involvement of the media and or intima. By consensus criteria, a pathologic diagnosis of IgG4-related aortitis/periaortitis required greater than 50 IgG4+ plasma cells/400× hpf, with more than 50% of the plasma cells staining for IgG4. The presence of storiform fibrosis, obliterative adventitial phlebitis, and eosinophil infiltration was also determined.2.5t test. Differences in the distribution of categorical variables (gender and signs/symptoms of vascular disease) were compared with the Chi-square test. A 2-sided P-value of <0.05 was considered statistically significant for all tests.Clinical variables were compared between the primary vasculitis and secondary IgG4-related vasculopathy groups. These included laboratory parameters, age at disease onset, gender, number of organs involved and signs/symptoms attributed to vascular involvement. Differences in continuous, nonparametric variables  were compared using the Wilcoxon signed-rank test while parametric variables (age and number of organs involved) were compared using the 3Among our cohort of 160 IgG4-RD patients, we identified large-vessel involvement in 36 (22.5%).3.13.1.1The demographic features of the large-vessel IgG4-RD patients are shown in Table 3.1.2We illustrate the spectrum of large-vessel disease in IgG4-RD by describing 6 representative patients in detail.3.2A 67-year-old man had a history of autoimmune pancreatitis, pericarditis, asthma, and peripheral eosinophilia of undetermined etiology. He had undergone a modified Whipple procedure for the erroneous diagnosis of pancreatic cancer before the correct diagnosis of lymphoplasmacytic sclerosing pancreatitis (now termed type 1 [IgG4-related] autoimmune pancreatitis) was established. He had become an insulin-requiring diabetic as a complication of this procedure. The patient was referred after developing swelling of the left parotid and bilateral lacrimal glands with associated left-sided proptosis.CT imaging confirmed the clinical findings of bilateral enlarged lacrimal glands as well as enlargement of the left lateral rectus muscle. Bilateral lacrimal gland biopsies yielded a lymphoplasmacytic and eosinophilic cellular infiltrate, lymphoid follicles with germinal centers, and prominent periductal sclerosis within the lacrimal glands. Culture, microbial staining, and flow cytometry were negative for infectious and malignant etiologies. Immunohistochemistry showed 50 to 100 IgG4+ plasma cells/hpf, with >90% of all plasma cells staining for IgG4. A diagnosis of IgG4-related dacryoadenitis was made. Because of the patient's diabetes mellitus, he was treated with rituximab (1000 mg × 2 doses). By 3 months, the patient's proptosis, parotid gland swelling, and long-term nasal congestion had resolved completely and his serum IgG4 concentration had decreased from 1310 to 226 mg/dL. Repeat imaging showed slight improvement of the left orbital findings, resolution of the right orbital findings, and improved paranasal sinus mucosal thickening. Five months following induction, proptosis returned and radiologic findings worsened, prompting another course of rituximab, followed again by clinical and radiologic improvement.Six months after the second rituximab course, the patient presented with recurrent episodes of right-sided amaurosis fugax. A magnetic resonance angiography (MRA) revealed moderate to severe stenosis of the right common carotid artery Fig. . A carot3.3A 54-year-old man with a history of non-Hodgkin lymphoma treated with rituximab + CHOP (cyclophosphamide/hydroxyurea/oncovin/prednisone) therapy 7 years before presentation experienced recurrent episodes of cardiogenic pulmonary edema, a non-ST segment myocardial infarction, and progressive kidney disease with proteinuria. Renal ultrasound examination revealed an incidental abdominal aortic aneurysm and bilateral common iliac artery aneurysms. A CT scan of the abdomen showed diffuse, nonatherosclerotic arterial disease with aneurysmal dilatation of the abdominal aorta, the common iliac arteries, and the superior mesenteric artery Fig. . A focalKidney biopsy revealed a combination of membranous nephropathy and chronic active interstitial nephritis with IgG4-dominant immune deposits along both the tubular and glomerular basement membranes. More than 20 IgG4-positive plasma cells/hpf were present within the interstitium. These features were diagnostic of IgG4-related kidney disease with both tubulointerstitial nephritis and membranous glomerulopathy. The kidney injury ultimately resulted in end-stage renal disease. The serum IgG4 concentration, 1980 mg/dL, was approximately 16 times higher than the upper limit of normal (reference 2–120 mg/dL). Other baseline immunologic parameters are shown in Table One month after beginning treatment, the patient developed unstable angina, prompting urgent coronary artery bypass grafting. Intraoperative findings included grossly visible aneurysmal left main and left anterior descending arteries, a whitish fibrinous discoloration of the ascending aorta without plaque on epiaortic ultrasound, and a whitish discoloration with friability of the left internal mammary artery. The left anterior descending artery was also noted to dissect easily. Operative specimens revealed a lymphocytic infiltrate of the aortic adventitia with scant plasma cells, the majority of which stained for IgG4.The patient was treated with rituximab (1 g times 2 doses) and was successfully tapered off prednisone in the following 3 months. Serum IgG4 had decreased to 435 mg/dL only 1 month after rituximab treatment, and the serum IgG1, C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), and C3 had all normalized. He was maintained on rituximab (1 g) every 6 months without disease progression or recurrence, but his end-stage renal disease was not reversible.3.4A 69-year-old-woman with a history of hypertension, asthma, and seasonal allergies presented with subacute dyspnea on exertion, palpitations, and vocal hoarseness. Physical examination was notable for a systolic bruit over the manubrium with radiation to the carotid arteries (right more than left). Echocardiogram revealed a mild left pulmonary artery stenosis with a right ventricular systolic pressure of 44 mm Hg. The cause of the pulmonary artery stenosis and hypertension was a 4 cm × 3 cm soft tissue mass encasing the left pulmonary artery on CT Fig. A. AnotheThe serum IgG4 concentration was elevated at 471 mg/dL . An assay for antineutrophil cytoplasmic antibodies was negative. The serum IgG4 was 286 mg/dL (2–120 mg/dL), IgE was 1390 IU/mL (0–100), ESR 38 mm/h (0–13 mm/h), and the serum CRP was 16 mg/L . Biopsy of the laryngeal mass demonstrated a lymphoplasmacytic infiltrate, focal areas of storiform fibrosis, scattered eosinophilic infiltrate, >100 IgG4+ plasma cells/hpf, and >40% IgG4+/IgG+ plasma cells. There was no evidence of multinucleated giant cells, granulomatous inflammation, leukocytoclastic vasculitis, or fibrinoid necrosis. She was treated with rituximab (1000 mg × 2 doses) with improvement in symptoms and declines in the serum concentrations of both IgG4 and IgE. Three months after rituximab, a repeat PET scan demonstrated decreased FDG avidity in the areas of previous involvement , 73 mm/h (0–13), and 155 mg/L , respectively. CT of the chest and abdomen showed mediastinal and hilar adenopathy, with the largest node measuring 2.4 cm in diameter. An excisional paratracheal lymph node biopsy from 2 years earlier was reviewed with additional immunostaining, revealing an IgG4/IgG ratio >50%.At follow-up 1 month after surgery, the serum IgG4 was 1148 mg/dL . He was started on prednisone 40 mg daily and tapered over 9 months. This treatment course was complicated by difficult to control diabetes, weight gain, and pneumonia. Upon tapering the prednisone to 10 mg daily, his serum IgG4 concentration and acute phase reactants began to rise. He was then treated with rituximab (1000 mg × 2 doses). At 2 months, the serum IgG4 concentration had declined from 590 to 363 mg/dL. He experienced another disease flare at 9 months and entered clinical remission with a normal serum IgG4 following a third rituximab course. Over the next several years, the patient was lost to follow-up for extended periods of time because of medical nonadherence. He ultimately died of complications of IgG4-RD following multiple hospitalizations for IgG4-related pleural disease. Autopsy showed active IgG4-RD in multiple organs including the pleura and retroperitoneum. Examination of the aorta demonstrated the infiltration of CD4+SLAMF7+ cytotoxic T cells, recently reported by our group in the context of IgG4-RD.3.6A 74-year-old-man with a history of prostate cancer, RPF causing hydronephrosis, and coronary artery disease presented with chest pain. During the evaluation for angina the patient was found to have an aneurysm of the ascending aorta, descending thoracic aorta, abdominal aorta, and bilateral iliac arteries. The ESR was 140 mm/h . He ultimately underwent 4-vessel coronary artery bypass and repair of the ascending aortic aneurysm, the pathology of which revealed “nonspecific” aortitis. The patient was treated with a tapering course of prednisone over 1 year, beginning at 40 mg/day. He was referred to our institution for consultation when his acute phase reactants began to rise again.The patient denied any active symptoms. Physical examination was revealing for bilateral iliac artery bruits. The serum IgG4 concentration was 287 mg/dL  and the serum IgG1 concentration 1070 mg/dL (340–890 mg/dL). The ESR and CRP were 90 mmg/h (0–13 mm/h) and 45 mg/L (0–8 mg/L), respectively. The patient's blood plasmablast concentration was elevated at 910/mL . Review of the patient's aorta pathology demonstrated a lymphoplasmacytic infiltrate involving the media and adventitia, storiform fibrosis, and >100 IgG4+ plasma cells/hpf. More than 50% of the IgG+ plasma cells expressed IgG4 . Imaging revealed bilateral hydronephrosis secondary to soft tissue thickening along the left pelvic wall as well as multiple, nonobstructive 2 to 3 mm ureteral and kidney stones. Bilateral ureteral stents were placed with resolution of acute kidney injury and symptoms. A PET-CT demonstrated intense FDG avidity of the pelvic wall mass, which extended inferiorly and encased the left iliac vessels and ureter. These findings were concerning for recurrent malignancy Fig. A and B.A CT-guided core biopsy was performed revealing a lymphoplasmacytic cellular infiltrate with normal flow cytometry and frequent IgG+ plasma cells, the majority of those being IgG4+ on immunohistochemical staining. No malignant cells were identified. These findings were consistent with IgG4-related RPF. The ESR was elevated at 77 mm/h (0–13 mm/h) while the CRP was nearly normal . The serum IgG4 was normal .The patient underwent treatment with rituximab. A follow-up PET/CT demonstrated substantial improvement in the size and FDG uptake of the left pelvic wall mass Fig. A and B. 3.8The clinical features of patients with primary IgG4-related vasculitis and secondary vasculopathy are compared in Table 3.8.1Thirteen patients (8% of the entire 160 patient IgG4-RD cohort) had primary involvement of their vasculature by IgG4-RD. Aortitis with or without aneurysmal formation, which affected 9 patients, was observed in the thoracic aorta in 8 patients, in the abdominal aorta in 4 patients, and in both the thoracic and abdominal aorta in 3 patients. Aortitis comprised the most common manifestation among those with primary IgG4-related large-vessel vasculitis, accounting for 69% of this group. Two patients (Patients 4 and 12) had aortic dissections and 1 (Patient 13) had a contained aortic perforation. All 3 of these aortic complications occurred in the context of aneurysm formation. Other manifestations of primary IgG4-related vasculitis included coronary aneurysms (Patients 3 and 5), carotid arteritis , pulmonary artery aneurysm (Patient 5), pulmonary arteritis (Patient 13), iliac arteritis with aneurysm formation (Patient 12), and iliac phlebitis (Patient 6). These are tabulated in Table 3.8.2Periaortitis in the setting of RPF accounted for 27 of 29 patients (93%) of secondary vascular involvement by IgG4-RD. Superior vena cava (SVC) compression (with impending SVC syndrome), renal artery stenosis, and pulmonary artery stenosis, with encasement of the relevant blood vessels by soft tissue masses, accounted for the remaining secondary manifestations. Four patients demonstrated evidence of both primary and secondary vascular involvement, 3 with RPF and 1 with a soft tissue mass encasing the pulmonary artery. Only 13% of patients with secondary vascular involvement presented with signs or symptoms directly attributable to their vascular involvement  had at least 1 and 54% (7 of 13) had more than 1 extravascular site of involvement. These included the orbit, ear/nose/throat areas, salivary glands, pericardium, lung, pancreas, lymph nodes, bile ducts, kidney, and retroperitoneum. Of the patients with aortitis, 2 of the 9 did not have any extravascular involvement. All patients with secondary vascular involvement had extravascular involvement by definition. RPF accounted for the majority of cases of extravascular involvement, but 11 (48%) had additional organ involvement separate from RPF.3.8.4Laboratory values at presentation and prior to the initiation of treatment are shown in Table 3.9Imaging studies were available on 32 of the 36 patients. Thirty patients (89%) had CT imaging available, of whom 20 (55%) had concomitant PET studies. Based on radiologic findings, the cohort consisted of 8 primary IgG4-related vasculitis patients, 19 secondary IgG4-related vascular involvement patients, and 5 patients with both primary and secondary findings.A total of 13 patients exhibited primary vascular findings. These findings are summarized in Table RPF was the most frequent CT finding among those with secondary IgG4-related vascular involvement. These patients consistently showed the pattern of an infrarenal, periaortic, soft tissue mass, located primarily in an anterolateral distribution and enhancing with contrast. Hydronephrosis was present in nine of the 25 patients (36%) with RPF and ranged in severity from mild to moderate. Two patients required ureteral stenting. Medialization of the ureters was also present in approximately half of patients with periaortitis Fig. . AdditioThree patients demonstrated secondary perivascular disease that did not involve RPF. One of these patients had a soft tissue mass encasing the superior vena cava; the second had a soft tissue mass encasing the left renal artery; and the third (Patient #7) had a soft tissue mass encasing and narrowing the main pulmonary artery. All 3 of these secondary lesions were associated with luminal stenosis of the involved vessel.3.10Diagnostic vascular pathology was available for 5 of the 13 patients with primary IgG4-related vasculitis Figs.  and 8. T3.113.11.1and had pre- and post-rituximab imaging available, 3 showed radiologic improvement, with reduction of the aortic wall thickening and enhancement  was complicated by friable, dissection-prone vascular tissue of the left internal mammary artery and left anterior descending artery during coronary artery bypass grafting. Only 1 patient was on glucocorticoids preoperatively. The aortic aneurysms in these cases were described as inflamed in appearance with white fibrinous discoloration and sometimes with adherent neighboring tissues. One aortic aneurysm showed intraoperative black areas of necrosis suspicious for impending rupture. All surgeries were successful in correcting the underlying vascular pathology and all of the surgical patients received rituximab following their IgG4-RD diagnosis.3.11.2and were on prednisone prior to induction therapy were successfully able to taper completely off of prednisone within 6 months of induction. The remaining 1 patient was lost to follow-up.Of the 23 patients in this subgroup, 1 was treated with tamoxifen plus prednisone, 6 with prednisone monotherapy, 13 with rituximab, 1 was observed without therapy, and 2 had no information about treatment available. The decision to use tamoxifen plus prednisone in the single patient was made by the referring rheumatologist prior to evaluation in our division. In general, prednisone monotherapy was used as initial treatment. The 7 patients not treated with rituximab responded to initial therapy both symptomatically and radiologically, obviating the need for treatment escalation. Eleven of the 13 secondary patients treated with rituximab had RPF with periaortitis as the vascular manifestation. The decision for rituximab in this subgroup was based on refractoriness to prior therapy (n = 6), baseline contraindication to glucocorticoids (n = 4), or discretion of the treating clinician (n = 3). Of those treated with rituximab, 5 showed radiologic improvement, 5 were unchanged, 1 worsened, and 2 did not have follow-up imaging available. The 1 patient that worsened despite rituximab was a patient with sclerosing mediastinitis and an enlarging mediastinal mass that encircled the SVC. Nine of the 13 rituximab-treated patients showed clinical improvement in their signs or symptoms, 3 were asymptomatic at treatment start date, and no follow-up data were available on 1 patient. Seven of the 8 patients who received rituximab 4IgG4-RD has been described in nearly every organ system in the body. Our series demonstrates that large blood vessels are also targeted in this disease and that this large-vessel involvement can be comprised of either a primary IgG4-related vasculitis or a secondary vasculopathy. These disease manifestations sometimes occur in the same patient. The primary vascular manifestations of IgG4-RD described in this series include large-vessel vasculitis of the aorta, pulmonary artery, iliac arteries or iliac veins, as well as a medium-vessel vasculitis involving the carotid or coronary arteries. The arterial beds affected by IgG4-related vasculitis are susceptible to aneurysm formation, which can occasionally be complicated by dissection or perforation. In contrast, arterial stenosis caused by primary IgG4-related vasculitis was not observed in our series, marking an important contrast with other large-vessel vasculitides, particularly giant cell arteritis and Takayasu arteritis. Some of our patients demonstrated aortitis as an isolated organ manifestation, but extravascular involvement typical of IgG4-RD was the norm in cases of primary IgG4-related vasculitis. the degree of avidity demonstrated in these patients could not be explained by atherosclerotic disease.The radiologic findings demonstrated in this cohort of patients were distinct from those generally found in atherosclerotic disease. Typical atherosclerotic aneurysm features include visible intravascular mural plaque with or without calcifications; diffuse, multifocal arterial involvement; and minimal if any vascular inflammatory changes in the absence of impending rupture. In contrast, imaging in this cohort demonstrated vessel wall thickening, wall enhancement by gadolinium, FDG uptake on PET studies, and aneurysmal dilatation. Although vascular wall inflammation is a recognized component of the pathogenesis of atherosclerosis both histologically and on PET imaging,,7,19,21 In contrast to the inflammatory infiltrate that centers upon the media in other forms of aortitis , the cellular infiltrate of IgG4-RD aortitis predominantly involves the adventitia, with lesser involvement of the media. This distinction has been made clear in a recent consensus statement on the surgical pathology of the aorta with a focus on inflammatory conditions. Similar to the pathology findings in the pancreas, major salivary glands, orbital tissues, and other organs affected by IgG4-RD, the histopathology and immunostaining of vascular involvement demonstrates a lymphoplasmacytic cellular infiltrate, storiform fibrosis, obliterative phlebitis, and an elevated ratio of IgG4/IgG-positive plasma cells.,21,22 These findings, which have been documented in both thoracic and abdominal aorta, are similar to those observed in our study. In addition, we describe the findings from a carotid resection with focal IgG4-RD involvement.The histopathology of large-vessel vasculitis in IgG4-RD has been described.The vascular system can also be affected in a secondary manner by the compressive effect of the tumefactive lesions for which IgG4-RD is known. In this cohort, these lesions occurred by far most often in the setting of RPF. IgG4-RD tends to encase the abdominal aorta beginning in the infra-renal region and extending caudally to the iliac arteries. Medialization of the ureters and hydronephrosis are common associated findings, leading frequently to permanent renal injury through postobstructive nephropathy. Patients with secondary IgG4-related vasculopathy appear to be at low risk for aneurysm formation compared to those with primary IgG4-related vasculitis. The abdominal aorta itself appeared to be minimally affected by the surrounding soft tissue in cases of RPF without any cases of aneurysm or stenosis. We did observe 3 cases of arterial narrowing as a result of secondary IgG4-related vasculopathy: one involving the superior vena cava, another the renal artery, and a third, the pulmonary artery.Perivascular soft tissue abnormalities are minimal in primary IgG4-related vasculitis, a condition in which the inflammation is centered on the vessel wall itself. In contrast, the predominant features of secondary IgG4-related vasculopathy are perivascular soft tissue enhancement and thickening with FDG avidity, but inflammation within the vessel wall itself appears to be minimal, at least as judged by radiology studies.,23 Whereas IgG4-related vasculitis predisposes to aneurysm formation and occasionally to dissection or perforation, secondary IgG4-related vasculopathy is more likely to cause arterial stenosis. The distinction between primary IgG4-related vasculitis and secondary vascular involvement is important in our evolving understanding of this condition, predicting outcomes, and guiding optimal therapy.Clinical differences also exist between these 2 categories of vascular disease in IgG4-RD. Patients with primary IgG4-related vasculitis have greater elevations in total serum IgG and trends toward greater elevations in IgG1, IgG4, and CRP. These patients are also much more likely to present with signs or symptoms directly attributable to the vascular involvement as opposed to those with secondary vasculopathy. Only 13% of patients with secondary vascular involvement presented with signs or symptoms directly attributed to their vascular involvement, a finding consistent with other reports. Such therapy is particularly beneficial in this aged population often with multiple other comorbidities that are exacerbated by glucocorticoids . Seven patients with IgG4-related vasculitis required surgical intervention while the nonsurgical patients treated with rituximab showed stable or improved vascular findings on follow-up imaging. Such findings underscore the importance of early identification and aggressive medical therapy in preventing surgical necessity and poor outcomes. Larger studies of patients with IgG4-related large-vessel disease, both primary and secondary, are required before definitive recommendations regarding treatment can be made.Although conclusions from this retrospective study with regard to treatment efficacy are necessarily circumscribed, IgG4-RD patients with vascular involvement appear to respond quite well to B cell depletion. Such treatment was effective for both primary vasculitis and secondary vasculopathy patients alike in regards to clinical improvement, steroid-sparing effect, and radiographic stability. Similar to the findings in this cohort, marked responsiveness of IgG4-RD to B cell depletion was recently published by our group in a prospective clinical trial involving 30 patients with IgG4-RD.Our data highlight the substantial frequency of vascular involvement in IgG4-RD, which affected 22.5% of our patient cohort. With nearly a quarter of IgG4-RD patients having some form of vascular involvement, clinicians should maintain a low threshold for obtaining vascular imaging in newly diagnosed patients in the interest of detecting such involvement early and preventing complications. In our experience, PET-CTA with the angiography component including noncontrast, postcontrast arterial and delayed phases is the most appropriate imaging study to evaluate the large vessel walls for evidence of enhancement, thickening, aneurysm, or FDG-uptake. This imaging modality is especially important in distinguishing true vessel wall involvement (primary IgG4-related vasculitis) from adjacent inflammatory tissue (secondary IgG4-related vascular involvement). In the event that vascular involvement is detected on imaging studies in IgG4-RD, the frequency of reimaging must be determined on a case-by-case basis, depending on the nature of the abnormalities found. IgG4-RD is typically an indolent disease by nature, however, and reimaging more frequently than every 6 to 12 months depending on the degree of blood vessel involvement is probably not indicated in most cases.Our study has both strengths and limitations. We used a large cohort of IgG4-RD patients from a single center. This cohort consisted of a wide variety of organ involvement, reflective of the systemic nature of IgG4-RD. The retrospective nature of this study is a limitation, which carries the potential for recall bias on the signs and symptoms attributable to vascular involvement. In addition, we may have underestimated the true frequency of vascular involvement in our cohort, because the entire cohort of 160 was not evaluated systemically by radiology studies.4.1IgG4-RD is a unique and treatable cause of primary large and medium-vessel vasculitis as well as secondary vasculopathy. Although the majority of patients with IgG4-related vasculitis are symptomatic at onset, this is a common disease manifestation with grave consequences that may warrant radiologic screening at the time of diagnosis. IgG4-related vasculitis usually takes the form of an aortitis with aneurysm formation. Periaortitis with relative sparing of the aortic wall is the most common type of secondary vascular involvement. IgG4-related vascular disease is associated with considerable morbidity and even mortality, but patients can be managed effectively if diagnosed early and treated appropriately.


To determine how important governmental, social, and economic factors are in driving antibiotic resistance compared to the factors usually considered the main driving factors—antibiotic usage and levels of economic development.A retrospective multivariate analysis of the variation of antibiotic resistance in Europe in terms of human antibiotic usage, private health care expenditure, tertiary education, the level of economic advancement (per capita GDP), and quality of governance (corruption). The model was estimated using a panel data set involving 7 common human bloodstream isolates and covering 28 European countries for the period 1998–2010.Only 28% of the total variation in antibiotic resistance among countries is attributable to variation in antibiotic usage. If time effects are included the explanatory power increases to 33%. However when the control of corruption indicator is included as an additional variable, 63% of the total variation in antibiotic resistance is now explained by the regression. The complete multivariate regression only accomplishes an additional 7% in terms of goodness of fit, indicating that corruption is the main socioeconomic factor that explains antibiotic resistance. The income level of a country appeared to have no effect on resistance rates in the multivariate analysis. The estimated impact of corruption was statistically significant (p< 0.01). The coefficient indicates that an improvement of one unit in the corruption indicator is associated with a reduction in antibiotic resistance by approximately 0.7 units. The estimated coefficient of private health expenditure showed that one unit reduction is associated with a 0.2 unit decrease in antibiotic resistance.These findings support the hypothesis that poor governance and corruption contributes to levels of antibiotic resistance and correlate better than antibiotic usage volumes with resistance rates. We conclude that addressing corruption and improving governance will lead to a reduction in antibiotic resistance. Escherichia coli and Staphylococcus aureus. People with infections caused by resistant isolates have much higher death rates as well as increased complications and suffering. Antimicrobial resistance also has important economic costs such as the use of more expensive drugs, waste and increasing the length of hospital stays [Antibiotic resistance is a growing international problem. This has led to increasing numbers of serious infections that are very difficult, or sometimes impossible to treat. Increasing resistance involves nearly all bacteria that infect people, including very common ones such as al stays –6.Antibiotic resistance develops and spreads wherever antibiotics are used, not only in medical facilities but also in the community. Poor infection control, poor water sanitation and poor hygiene all facilitate the spread of resistant bacteria from person to person. The majority of antibiotic usage worldwide is in food animals –9. This The general perception of antibiotic resistance is that it is almost entirely related to the amounts of antibiotics used, not only in the broad sense of comparative usage by different countries but also in individuals –17. HoweWe postulate that a fuller understanding of antibiotic resistance requires a unified approach which combines not only the amount of antibiotic usage but also other factors that may impact on the quality of antibiotic usage, control and the spread of resistant bacteria at national levels. We therefore undertook a multivariate analysis of determinants of antibiotic resistance in bacteria causing bloodstream infections throughout Europe.For the empirical analysis, we used a newly-constructed panel data set covering 28 countries in Europe over the period 1998–2010  (+),GOV the quality of governance, defined as the control of corruption in society measured on a discrete scale from zero to six (-),PHE private health expenditure as a percentage of national income (+),PGDP per capita gross domestic product in constant (2000) price  (-),TED percentage of tertiary educated people in total population (-),AGR percentage of people employed in the agriculture sector out of total employment (+)i Country-specific and time-invariant fixed factors.μt Time Effects (i.e. binary variables for each year except for the first year) to capture time varying common shocks or global trends, andγi,t is the idiosyncratic error term that captures all other determinants of ABRεABU and GOV are the main explanatory variables that we are interested in comparing as determinants of ABR. ABU is the variable commonly associated with antibiotic resistance and which is assumed to have a positive impact on antibiotic resistance. GOV captures the degree to with the rule of law and social norms are honored in a given country. This will not only affect the operations of government, infrastructure and social services but also the private sectors and the quality of medical practices. The other variables are the control variables which are simultaneously correlated with the main explanatory variables of interest as well as the dependent variables. It is important to include these variables in the model to mitigate omitted variable bias. PHE may have a positive impact on antibiotic resistance if greater access to private medical sector leads to increased antibiotic usage may have a negative impact on antibiotic resistance, if economic prosperity is positively related with greater public understanding of health risk, better housing and on the availability of better public infrastructure. TED would have a negative impact on antibiotic resistance if higher education levels result in greater public understanding of antimicrobial-resistance health risks.European Centre for Disease Prevention and Control [Europe was chosen for estimating the model because it is the only region where good data for all of our parameters are available across multiple countries. Our panel data set covered 28 countries in Europe over the period 1998–2010. The countries covered are Austria, Belgium, Bulgaria, Cyprus, Czech Republic, Denmark, Estonia, Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Slovakia, Slovenia, Spain, Sweden, and United Kingdom. The country- and time-coverage of the analysis are dictated by the nature of availability of data on the dependent variables (ABR). These data on antimicrobial resistance were compiled from EARS-Net Database of the  Control . This daIn this study, we looked at 25 pathogen/antibiotic combinations which are grouped under seven pathogen classes. The pathogen classes and the relevant antibiotics are listed as follows:Streptococcus pneumoniae resistance to penicillins (PNSP) and macrolides (MNSP)S. aureus resistance to methicillin (MRSA) and rifampicinPseudomonas aeruginosa resistance to amikacin, aminoglycosides, carbapenems, ceftazidime, fluoroquinolones and piperacillin/tazobactamKlebsiella pneumoniae resistance to 3rd gen cephalosporins, aminoglycosides, carbapenems and fluoroquinolones.E. coli resistance to 3rd gen cephalosporins, aminoglycosides, aminopenicillins, carbapenems and fluoroquinolones.Enterococcus faecium resistance to aminopenicillins, high-level gentamicin and vancomycinEnterococcus faecalis resistance to aminopenicillins, high-level gentamicin and vancomycinpneumoniae’ group of resistances are not available for Greece, while data for ‘P. aeruginosa’ and ‘K. pneumoniae’ are not available for Slovakia. After allowing for these data gaps, the regression estimates are based on 161 to 298 observations.Data for ‘S. European Surveillance of Antimicrobial Consumption (ESAC) Yearbook 2009 [World Development Indicators database of the World Bank [Data on antibiotic usage are from ook 2009 . In thisrld Bank .PGDP is per capita gross domestic product (GDP) converted to constant year 2000 international dollars using purchasing power parity rates. Tertiary school enrolment is measured in terms of the ratio of total enrolment in tertiary educational institutions, regardless of age, to the population of the age group that officially corresponds to the level of education shown. Private health care expenditure (PHE) includes direct household (out-of-pocket) spending, private insurance, charitable donations, and direct service payments by private corporations. We use this as a percentage of total Gross Domestic Product in order to normalise for the size of the economy. Agriculture share of employment (AGR) is based on both public and private employment in agriculture (which corresponds to ISIC revision 2), and receive any kind of a remuneration. These variable definitions are from World Development Indicators [dicators .In this paper, we specifically focus on the quality of governance in comparison to antibiotic usage as determinants of antibiotic resistance, while controlling for the other possible determinants. We measure the quality of governance in terms of the control of corruption, which is the most ubiquitous manifestation of governance quality in policy debates –24. HencGOV) comes from the International Country Risk Guide developed by the Political Risk Services Group [The indicator governance (es Group . This ines Group  The resuDDD (for Netherland in 2004) to a maximum of 45 DDD per day per 1000 inhabitants (for Greece in 2007). According to the country-level data (not reported here for brevity but available in i). However, ignoring country-specific fixed effects could cause omitted variable bias, because country-specific and time-invariant factors might be correlated with both ABR and the explanatory set. These factors include geography, historical experience, legal origin, ethno linguistic fragmentation and culture. We therefore re-estimated the model using the fixed effects (FE) technique. This technique involves estimating the equation after demeaning the variables to purge possible country-specific fixed effects in order to isolate the ‘within country’ relationship.The model is initially estimated using the Pooled Ordinary Least Squares (POLS) method, which estimates the equation excluding the country—specific fixed factors  may suffer from a measurement error. Both types of endogeneity lead to biased and inconsistent estimates for the coefficients. When the explanatory variables are potentially endogenous, we need to be cautious in inferring causality from the estimated coefficients.We therefore use the system Generalized Method of Moments (system GMM) technique as a robustness check. This technique consistently estimates the equation even in the presence of endogenous explanatory variables and measurement error, and thus allows us to be more confident in inferring causality –31. The System GMM generates a set of internal instruments for the right hand side variables which are uncorrelated with the error term. This set consists of the appropriate number of lags of the right hand side variables as instruments in the differenced version of the original equation and the first differences (and the appropriate number of lagged first differences) as instruments in the original level equation. The validity of the instrument set can be tested through a Hansen test of over-identifying restrictions. However, this test is weakened if the number of instruments is large relative to the number of observations, and works best if the number of time periods in the panel is small . TherefoABU is measured in DDDs; ABR, PHE and TED are percentages; and GOV is an index (0 to 6) and PGDP is in thousand dollars (expressed in logs). Therefore, the regression coefficients are not directly comparable. We therefore report results using the beta coefficients to facilitate comparing the relative impact of each explanatory variable on the dependent variable. The beta coefficient places all regression coefficients on a comparable basis by measuring the impact of change in a given explanatory variable on the dependent variable, using the standard deviation as the common denominator.Note that the unit of measurement used varies among the variables employed in our analysis: 2 = 0.28) is attributable to variation in antibiotic usage in people. When the time effects are also included to take into account possible shocks or global trends (not captured by the other explanatory variables) impacting antibiotic resistance uniformly across all countries, the regression explains 33 percent of the total variation in antibiotic resistance. Once the control of corruption indicator is included as an additional explanatory variable, 63 percent of the total variation in antibiotic usage is explained by the regression. The complete multivariate regression only accomplishes an additional 7 percentage point in terms of goodness of fit, indicating that corruption is the main socioeconomic factor that explains antibiotic resistance. The income level of a country appeared to have no effect on resistance rates when we also controlled for the quality of governance in the multivariate analysis.Our results support the crucial role of governance in determining antibiotic resistance across these countries. When we use the simplest of our estimations, only 28 percent of the total variation in antibiotic resistance  are each associated with a reduction of 0.2 units in ABR.Focusing on the complete multivariate regression (Column 3), the coefficients of all variables have the expected sign and, except that of the employment share of agriculture, are statistically significant at the one percent or five percent level. In terms of magnitudes of the coefficients, the governance variable has the greatest association with antibiotic resistance; a one unit improvement in the indicator is estimated to lower antibiotic resistance by 0.4 units. A similar reduction in antibiotic usage only reduces resistance by 0.3 units. As for the other variables which are statistically significant, a one unit increase in growth in tertiary enrollment brings about a reduction of 0.1 units in 2: adding additional variables into the specification practically makes no difference, indicating the central role of fixed country characteristics such as culture and geography. Interestingly, when fixed country characteristics are controlled for, governance is still a statistically significant determinant of antibiotic resistance. The estimated coefficients suggest that, in the within country context, a one unit improvement in the corruption score is associated with a 0.2 unit reduction in ABR.The FE results reported in Columns –6, howevThe system GMM estimates are given in Columns 7–9. The results suggest a causal relationship between our variables of interest after addressing all possible sources of endogeneity. The diagnostics indicate that the model has been adequately estimated. The p-value of the Hansen test of over-identifying restrictions indicates that the instrument set, which identifies the exogenous variation in the explanatory variables, is valid. Moreover, the p-value is not unrealistically high, which would have suggested an over-fitting bias. The p-value of the AR (2) test also indicates that there is no second order serial correlation in the residuals, a necessary condition for consistent estimation. An unrealistically high p-value is a symptom of instrument proliferation in which case the Hansen test is weakened .ABU is estimated with precision only in the second regression, and even then it is significantly smaller in magnitude compared to the coefficient of GOV . In the complete multivariate model (Column 9), the coefficient of ABU is not statistically significant even at the 10% level. In contrast, the estimated impact of GOV is statistically significant at the one percent level, and the coefficient indicates that an improvement in the “control of corruption” indicator by one unit reduces antibiotic resistance by approximately 0.7 units. Interestingly the causal impact of private health expenditure is also estimated with precision: a one unit reduction reduces ABR by 0.2 units. The system GMM results also suggest that antibiotic resistance in the previous period does not affect present resistance, and thus ABR is not a persistent variable.Control of corruption remains the most important determinant of antibiotic resistance according to the system GMM results. The coefficient of The results of our empirical analysis show that factors other than antibiotic usage are potentially very important in explaining the different levels of resistant bacteria seen in different countries Figs.  and 2. TPrivate health expenditure was also an important factor. The higher the percentage of private health expenditure in a country, the larger was the degree of antibiotic resistance. The reason for this is not clear. We postulate that when healthcare is being delivered predominantly in the private sector, there are less controls and supervision of what is being done. This then may mean that there are fewer controls on broad-spectrum agents, the length of time of drug therapy and the volumes used.We postulate that when Quality of Governance is poor, then there are likely to be less effective controls of antibiotic use . Thus, not only will more antibiotic resistant bacteria develop but the spread of these resistant bacteria will also be easier. This is because there will be less supervision and enforcement of laws that cover issues, not only of human medicine, but also of food and water safety. Reducing antimicrobial resistance requires a policy mix aimed at lowering antibiotic usage in people and, perhaps even more importantly, developing better controls on corruption.Surprisingly, our results suggest that antibiotic resistance in the previous period does not affect present resistance. This implies that previously high levels of antimicrobial resistance in a country are not insurmountable barriers to future improvements.The data we have shown only pertain to Europe. This is because this is only region where good data are available from multiple countries. Europe has data not only on the issues of government effectiveness and rule of law etc, but more importantly on the magnitude and extent of antimicrobial usage and resistance levels in serious bloodstream infections caused by common bacteria such as S. aureus and E. coli. We suspect that these same issues will pertain worldwide (e.g. corruption). In developing countries, they are likely to be much more of a factor, and quality of governance will likely explain why antibiotic resistance is so much higher in these countries than in most countries in Europe. The fact that these data are publicly available for Europe, suggest that issues of governance are also much better in Europe than in most of the rest of the world.To our knowledge, this is the first cross-national examination of the determinants of antibiotic resistance. Our results, based on a systematic econometric analysis using a panel data set consisting of 29 European countries over the time period 1998–2010, suggest that governance is an even more important determinant of antibiotic resistance than antibiotic usage in people. The estimated impact of governance is large and highly statistically significant, using state-of-the-art econometric methodologies that pay particular attention to issues of endogeneity. The results are robust to the use of alternative estimation methods.GOV) is based on subjective assessment by experts. This variable comes from the International Country Risk Guide developed by the Political Risk Services Group [There are, however, limitations in our study. One of our main explanatory variables, “quality of governance”  is the fundamental determinant of economic growth—our results suggest that improving governance could be similarly fundamental in confronting the issue of antibiotic resistance. Improved governance at the national level is likely to imply better practices in the health sector, including controls and supervision of antibiotic use. It is also likely to lead to healthier outcomes in a whole array of other related areas that are intertwined with the issue of antibiotic resistance, such as the agriculture sector, food and water safety.S1 Table(XLSX)Click here for additional data file.


In the injured spinal cord, chondroitin sulfate proteoglycans (CSPGs) are the principal responsible of axon growth inhibition and they contribute to regenerative failure, promoting glial scar formation. Chondroitinase ABC (chABC) is known for being able to digest proteoglycans, thus degrading glial scar and favoring axonal regrowth. However, its classic administration is invasive, infection-prone and clinically problematic. An agarose-carbomer (AC1) hydrogel, already used in SCI repair strategies, was here investigated as a delivery system capable of an effective chABC administration: the material ability to include chABC within its pores and the possibility to be injected into the target tissue were firstly proved. Subsequently, release kinetic and the maintenance of enzymatic activity were positively assessed: AC1 hydrogel was thus confirmed to be a feasible tool for chABC delivery and a promising device for spinal cord injury topic repair strategies. SCIestigate ,5. It isestigate . The iniestigate .In vivo studies provided key evidences that chondroitinase ABC (chABC), a bacterial enzyme able to digest chondroitin sulfate glycosaminoglycans (CS-GAGs) of CSPGs, fosters axonal regeneration and sprouting, promotes plasticity of uninjured pathways and, most relevantly, supports functional recovery in various animal models [l models ,7,8,9. Hl models ,7. Thesel models ,11. Amonl models ,13. In this study an injectable hydrogel from agarose and Carbomer 974P, here briefly named agarose-carbomer (AC) in accordance with previous works ,15,16, wIn vitro, in vivo and ex vivo release studies showed an independent combination of fast diffusion-controlled kinetics for smaller molecules together with slow diffusion-controlled kinetics for larger ones. Moreover, a preserved functionality of loaded substances was always achieved, confirming the absence of any chemically stable interactions between gel matrix and loaded molecules.Here, release tests were first performed with Texas-Red dextran (Tx) to confirm the possibility of achieving an effectively deliver of chABC-like molecules ,19. Delii.e., being capable of maintaining the activity of the carried substance: chABC activity was here specifically assessed through its efficacy of digesting decorin, a small proteoglycan [A key quality for a delivery system is to be neutral, eoglycan .Carbomer 974P (a highly branched polyacrylic acid) was provided by Fagron , high purity sodium hydroxide (NaOH) was purchased by Fluka , and agarose was provided by Invitrogen . The solvent used was PBS , purchased by Sigma-Aldrich .AC1 hydrogel in according with previous studies [Hydrogels were synthesized by bulk reaction in PBS between agarose and Carbomer . To preserve the actual morphology of the hydrogel under complete swelling, freeze-drying (24 h) was applied to remove all the liquid phase by sublimation. Because of the low operating values of temperature and pressure of this procedure, gel polymer chains retain the same conformation they show in wet conditions ,22.AC1 hydrogel were performed at 25 °C using a Rheometric Scientific ARES instrumentation  equipped with 30 mm parallel plates, with a 4 mm gap between them [The storage moduli and time sweep of een them . OscillaTx, being of about 70 kDa molecular weight , was chosen because of its hydrodynamic radius similar to chABC ,19 and f25 μL chABC , with a concentration of 2 U/0.5 mL in PBS, was mixed with 25 μL of hydrogel, once gel had already been irradiated and cooled down to 37 °C. chABC remained homogeneously entrapped in the pores of the hydrogel at the end of gelation process, similarly to what happened inside Tx-loaded hydrogels. Every well containing a hydrogel was filled with 100 μL of distilled water and stored at room temperature. Aliquots of 100 μL were collected from every gel samples at defined time points , filling wells with fresh solution. Subsequently, 10 μL of decorin 2 mg/mL  was added to 40μL of each samples and enzymatic digestion was allowed to proceed for 12 hours at 37 °C. The resulting products were analyzed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), in order to assess any potential adverse effects of the hydrogel on chABC enzymatic activity. Solutions of pure decorin and decorin digested by chABC were used as controls during SDS-PAGE analysis.Where applicable, experimental data were analyzed using Analysis of Variance (ANOVA). Statistical significance was set to p value <0.05. Results are presented as mean value ± standard deviation .The investigated hydrogel is a chemical gel synthesized through a statistical block polycondensation between Carbomer 974P A and agaIndeed, Carbomer 974 P carboxylic groups constitute the main cross-linking sites to be reacted with hydroxyl groups from agarose, altogether giving rise to the three dimensional matrix. These polymers were selected because of their well-known biocompatibility in central nervous system (CNS). Agarose has been widely used in biomaterials for spinal cord injury repair strategies , while cThe G' and G'' moduli trend versus oscillation stress is reported in critical stress) the viscosity drops drastically because the material collapses and gel-sol transition occurs. Outside the LVR, G' trend begins to decrease, while G'' values remains almost constant, indicating that the viscosity of the material increases with respect to its elasticity. At the crossover point .As hydrogel injectability was tested confirming previous results , its abiplateau). Moreover, it has to be noted that all loaded Tx was released, underlining not only the absence of any stable bonds between hydrogel polymeric network and loaded Tx but also hydrogel capacity to deliver even those molecules deeply entrapped in its core.i.e., in this specific case chABC enzymatic activity. SDS-PAGE plot reported in Nevertheless, besides being able to transport molecules with high hydrodynamic radius, a delivering hydrogel must preserve drug biochemical properties, As can be noticed, each time point  is described by a lane (a–d) with an evident band in the range of 37–50 kDa. Specifically, this band is close to 45 kDa, which represents the molecular weight of decorin main digestion product (lane e). Indeed, decorin is known to consist of two or three asparagine-bound oligosaccharides, weighting about 45 kDa each, and a glycosaminoglycan chain. After chABC digestion just its 45 kDa core is still preserved ,29. ThesHence, the absolute neutrality of agarose-carbomer hydrogel was proven with respect to chABC biochemical features for the whole length of the delivery interval.Up-to-date spinal cord repair strategies cannot be considered separately from degrading the glial scar, which is the main obstacle to axonal regrowth. chABC has been proven to successfully digest CSPGs, but it presents remarkable administration shortcomings. In this study, a promising agarose-carbomer hydrogel was investigated, demonstrating its capacity to release molecules with high hydrodynamic radius over a one-week period. The material showed the ability to entrap chABC molecules and exhibited coherent mechanical properties to be easily injected into the target tissue. Furthermore, hydrogel loading did not induce any stable bonding with chABC nor any chABC denaturation: an effective digestion of scar-like molecules, such as decorin, was observed throughout the whole expected delivery timeframe. Agar-carbomer hydrogel was thus confirmed to be a potential delivery tool for chABC in spinal cord injury repair strategies.


On knowledge scale, respondents had a mean score of 8.0 ± 1.7, attitude 5.8 ± 3.4, perception 23.4 ± 4.1, misconceptions 8.0 ± 2.1 and views 8.0 ± 3.9. The respondents mean score was 53.2 ± 9.1 (64.9 %). Overall, level of education, marital status, religious beliefs, and employment status has significant (p < 0.001) associations with KAP-MV. Knowledge was significantly correlated with respondents’ attitude , perception , and views . Family members of PLWHA with less knowledge scorehad more negative attitude, perception and views. Level of education, marital status, religious beliefs and employment status were identified as key barriers. Interventions targeting family members of PLWHA are warranted. Practice implications are as follows: Encourage role of family members.Deploy interventions.Minimize barriers.Change misconceptions.We intended to assess knowledge, attitude, perception, misconception and views (KAP-MV) of family members of PLWHA. A cross-sectional retrospective study conducted in Anti-retroviral centre of Mahatma Gandhi Memorial—MGM hospital, Warangal, Telangana, South-India from July to September 2014. A questionnaire containing 41 items was distributed among adult family members accompanying patients living with HIV/AIDS-PLWHA. Level of KAP-MV was categorized into poor (0–28), average (29–55) and good (56–82). Analysis was performed by Pearson’s Chi square, analysis of variance and Spearman’s correlation test on 41 variables using SPSS version 21 and  Verging on forth decade of acquired immune deficiency syndrome-AIDS epidemics, India ranks the third largest human immune virus (HIV) epidemics in the world  directly or indirectly face embarrassment, shame, ostracism and stigma from the community,  institute of Medical Sciences, Warangal, Telangana state.We intended to assess the knowledge, attitude, perception, misconception and views (KAP-MV) of adult family members of PLWHA in South India. The study aim was to delineate the predictors that affect adult family member’s KAP-MV.We have objectively assessed the KAP-MV of adult family members within a cohort of HIV/AIDS positive patients. The assessment of general knowledge has identified the level of understanding of adult family members towards HIV/AIDS, attitudes have addressed their feelings, perceptions have addressed the possible causes of HIV/AIDS, misconception have investigated their ideas towards certain myth about PLWHA and views examined certain daily contact activity of measures to prevent HIV transmissions.A cross-sectional survey was conducted using self-administered questionnaire "" among aThe population was composed of all adult visitors accompanying patients with HIV/AIDS visiting the main hospital. PLWHA adult family members were informed about the study prior to their participation. Those who agreed to participate were consented.An online sample size calculator “Creative research systems” , not willing to participate and not able to provide written consent were all excluded from the study.We have developed structured study survey-questionnaire which was self-administered to collect socio-demographics and the five domains outcome measures (KAP-MV). The questionnaire items used in this study was based on the World Health Organization (WHO) AIDS program knowledge, attitudes, beliefs and practices (KABP) survey in 1990, . It was divided into six parts. Part A related to respondent’s socio-demographic background (9 items), Part B was relevant to knowledge regarding HIV/AIDS (5 statements), Part C on risk perception towards possible causes of HIV/AIDS (12 statements), Part D on AIDS attitude scale (5 statements), Part E on respondents views about measure to prevent HIV transmissions (12 statements) and Part F on misconceptions towards HIV/AIDS (5 statements).DKB] on daily bases from July to September 2014.All the questions were translated from English to Telugu (back and forward method) by expert faculty members (number of 6). The Telugu version was used for the study participants. The questionnaire was tested for reliability, psychometric and internal validity. The internal consistency estimate of reliability of test scores (Cronbach’s Alfa) was found to be 0.76 indicating a good construct of KAP-MV. Furthermore, KAP-MV was piloted and post piloting phase the questionnaire was modified to meet the compatibility of local settings and suit the study participants. The results of the piloting phase were not included herein. The questionnaires were completed in the waiting room and took an average of 30 min to complete. The completed questionnaires were collected and retrieved by one investigator . More than half of respondents (55.1 %) were 18–24 years of age, had attained the level of higher educational degree or above (63.9 %), and single (59.6 %). More than half of respondents (58.4 %) were students, and have faithfully believed in God the Almighty (78.3 %). The mean income was 25,000 ± 2000  Indian Rubies-INR and mean number of adult family members living at home with their PLWHIA was 4.5 ± 1.3. The characteristics of the respondents were presented in Table The total mean scores of the study sample on respondent’s knowledge, attitude, perception, views and misconception (KAP-MV score) was found to be 64.9 ± 9.1. Which was categorized good (reference ranging from 56 to 82). Table The adult family members’ feeling towards PLWHA was presented in Table The sum of perception scores can range between 0 and 28 and respondents overall mean score in this domain was 23.4 ± 4.1. The analyses of data have shown that in most statements concerning perception and beliefs about the possible causes of HIV infections, the majority of respondents had good positive perception regarding HIV causes. However, some questions showed less perception towards causes of HIV/AIDS transmission. For example, only 337 (65.4 %) and 147 (28.5 %) knew that kissing and tattoos cannot transmit HIV-infections, respectively, [Table Overall respondent views towards preventing HIV transmission mean score was 21.0 ± 3.9 (reference range from 0 to 24). Generally, the views of adult family members were highly regarded as good towards certain issues concerning PLWHA and HIV/AIDS as a transmitted disease. For instance when respondents were asked that they avoid taking illicit drugs/use of intravenous drugs, avoiding sharing needles and syringes, having sex with only one faithful uninfected partner, using condoms during sexual intercourse and treating sexually transmitted diseases (STDs) promptly with more than 90 % had have answered correctly. On the other hand less than 90 % but more than 80 % responded correctly to the followings: screening donated blood before transfusion, not sharing food with an infected person, do not stay with infected person on same house and do not have casual contact with infected person. Furthermore, less than 80 % and over 70 % of respondents have shown correct answers to preventive measures as not sharing toilets with an infected person, isolating people living with HIV/AIDS (negatively worded) and avoid mosquito bites for HIV transmission, [Table The majority of the respondents had less misconception about HIV/AIDS, with 65–90 % correctly answering the five statements. However, many misconceptions were still noted relating to HIV/AIDS, with 34.6 % of respondents believing that love could be a reason for HIV/AIDS and 27.6 % believing that AIDS is a punishment from the God, [Table p < 0.001) and religion (p < 0.001). Knowledge was significantly correlated with attitude , views , perceptions  and total KAP-MV  towards HIV/AIDS. Adult family members of PLWHA with less knowledge score had more negative attitude, perception and views towards preventing HIV infections. Furthermore, misconceptions were highly associated with PLWHA families with negative views  and less perceptions . Age, level of education, marital status, religion status and religious beliefs were significantly associated with most attitude scores, (p < 0.001) and were minimally associated with respondents perception about HIV/AIDS, (p < 0.001). Results have revealed that all the sociodemographic variables were significantly associated with respondents’ views, (p < 0.001). Religious status (p < 0.001) was significantly associated with respondents’ misconceptions. The results of the correlations were illustrated in Table The results shown in Table The main finding of the current study was the satisfactory outcome of the KAP-MV survey in term of evaluating the adult family member basic knowledge, attitude, perception, misconception and views of their beloved PLWHA. We have used a multiple component questionnaire with diverse domains to the level of understanding of adult family members towards HIV/AIDS, feelings, perception about possible causes of HIV/AIDS, misconception towards certain myth about PLWHA and views on certain measures to prevent HIV transmissions. The instrument used in our study demonstrated clarity to communicate the intended meaning, coherency to define the logic wording, extendibility to accommodate newly added items with the domains, unbiased to allow for applicability and complex to meet the desired study purposes Table .Table 4SMany studies has used the WHO-KABP scale to assess the knowledge, attitude and practice towards PLWHA from past decades and applied on different groups, subgroups and geographical populations. To the best of our knowledge, this is the first study focused on PLWHA adult family members KAP-MV residing in high prevalent Warangal district, South-Central India.Nearly two-third of respondents have had a clear knowledge about the abbreviation used for HIV/AIDS. The adult family members of PLWHA had expressed a positive answer towards HIV/AIDS and high level of KAP-MV score on issues related to HIV/AIDS transmission, considering the majority of the respondents had answered correctly. Also the notation that AIDS is a hereditary disease or AIDS is a cured disease and that there is a vaccine for AIDS were equally answered correctly by respondents in a very high percent. However, a low percent of the respondents believed that AIDS is a hereditary disease and that there is an active treatment to cure AIDS, which was consistently very low compared with other results conducted in various populations,  . For example, 28.7 % of the respondents opinioned that PLWHA should be isolated from the community or special centre, avoiding mosquito bites to prevent HIV transmission (21.2 %) and unwillingness to share toilets with HIV infected people. Furthermore, when taking into consideration that the majority of the respondents had a positive views towards PLWHA and the level of opinions about PLWHA were above average for this attitude may be that few of adult family members do not have much positive opinion or views towards people with AIDS.Our study demonstrated that in general average number of the respondents harbored misconceptions. For instance, 34.5 % of the adult family members agreed with the statement that love is a reason for HIV/AIDS and AIDS is a punishment from God (27.6 %). In parallel, this proportion was slightly higher than in Ayranci and co-workers study (23.2 %),  of KAP-MV towards HIV/AIDS high prevalent in Warangal district, South India. The level of education, marital status, religious beliefs, and employment status has been identified as key barriers for adult family members caring for PLWHA. Interventions targeting adult family members of PLWHA are warranted to raise their awareness and knowledge, improves attitudes, minimize the negative views and misconception for more proactive role.Encourage the role of family members for supporting PLWHA.Deploy interventions to raise the knowledge, attitudes, perception and views of family members of PLWHA.Minimize the barriers for adult family members caring for PLWHA.Change the family members misconceptions about PLWHA.The following are highlighted as implications of the study on clinical practice:


IgG4-related disease was first reported in 2001 and was officially named in 2010. It is now considered as a systemic disease that might affect any organ system. The characteristic pathological changes of IgG4-related disease are extensive infiltration of IgG4-positive plasma cells. IgG4-related disease is a kind of benign lesions, but there has not been well-defined standard treatment so far. Patients usually respond well to corticosteroids. The prognosis of IgG4-related disease is perhaps good as long as early detection and treatment.We report one case of IgG4-related disease with a 16-years anamnesis with multi-pseudotumor masses. He was diagnosed with chronic kidney disease with wide interstitial renal fibrosis. And he received glucocorticoids therapy. After 2 month therapy, the serum creatinine, erythrocyte sedimentation rate, and serum IgG4 decreased significantly. The discussion includes presentation, clinical course, diagnosis, and prognosis of IgG4-related disease. The case and discussion highlight the importance of diagnosis and the good prognosis of IgG4-related diseases.Our case highlights the importance of diagnosis and the good prognosis of IgG4-related diseases. IgG4-related disease is a systemic fibro-inflammatory immune-mediated disorder and now recognized in almost every major organs. Characteristics of the disease is multiple lymph nodes and the response to glucocorticoids therapy is well. In such case, he had a history of 16 years with multi-pseudotumor masses and misdiagnosed for 16 years, if the doctors were not awareness of higher serum immunoglobulin G4 (IgG4) than normal, the correct diagnosis may be missed or delayed. Consequently, appropriate treatment for IgG4-related disease would also be delayed or not provided and likely result in increased morbidity and mortality.IgG4-related disease is a systemic fibro-inflammatory immune-mediated disorder and progresses slowly. In the present patient the course of IgG4-related disease appears to be benign. The prognosis of IgG4-related disease depend on early diagnosis and treatment. IgG4-related disease is now considered as a systemic disease that might affect any organ system, including kidneys, lymph nodes, and thyroid gland, with progressively growing fibro-inflammatory lesions causing a mass effect. Diagnostic criteria were established as: IgG4 plasma level of >135 mg/dl and an IgG4/IgG plasma cell ratio of >40 % with >10 and IgG4-positive plasma cells per HPF. Macroscopically, these diseases cause diffuse organ swelling and formation of pseudotumor masses. Patients usually respond well to corticosteroids, but highly active diseases may require other immunosuppressive therapies. IgG4-related disease was first reported in 2001, which came from autoimmune pancreatitis (AIP) and was officially named in 2010. The prognosis of the disease has not been clearly defined. We hereby reported a 54-year-old male with IgG4-related disease, a history of 16 years with multi-pseudotumor masses, he was diagnosed as chronic kidney disease with Scr 545 µmol/L and interstitial renal fibrosis widely. And the response to glucocorticoids therapy was well. After 1 month therapy, the Serum creatinine (Scr), erythrocyte sedimentation rate (ESR), and IgG4 decreased significantly.IgG4-related disease was diagnosed in a 54-year-old male with lumps in both orbital cavity for more than 16 years and lymphadenopathy in mediastinum for 11 years prior to hospital admission. In March 2000, a 54-year-old man was present in a local hospital because of exophthalmos for one year. The physical examination showed that there was hard lumps out of the top of left orbital cavity (0.5 × 0.5 cm) and larger tender bilateral submandibular lymph nodes. A computed tomography (CT) scanning and ophthalmic ultrasound indicated that occupying lesion located in left orbital cavity. Scr was 76 µmol/L. Pathology of lumps in the orbit was “inflammatory pseudotumor (benign lymphoma)”, and he was given prednisone, 30 mg qd. After 2 weeks, bilateral submandibular lymph nodes shrunk significantly (0.3 × 0.3 cm).During April 2000 to May 2004, the patient did not go to see any doctor for the above disease. At June 2004, he received CT scanning examination, and there still was swelling lymph nodes in mediastinum and abdomen. He was given prednisone  and proteinuria was positive (+), then he accepted some relative treatments. But at early of May 2015, Scr increased to 488 µmol/L, so he was admitted to our hospital. His prior medical history also included mild lower extremity numbness for more than 5 years.2+ and P3+ were normal. Anti-glomerular basement membrane antibody and anti-neutrophil antibody was negative. The ratio of light chain κ/λ in urine and blood were normal. Urinary albumin-creatinine ratio (ACR) was 0.25 g/gCr. The serum immunoglobulin G (IgG) was 57,500 mg/L  accounting for 50.3 % of total immunoglobulin, significantly higher than normal . Serum immunofixation electrophoresis was negative. Level of C3 and C4 were normal.Results of a physical examination revealed that there were enlarged lymph nodes behind the left ear, right groin and left orbital cavity, each was about 1.0 × 1.0 cm. There was no malar rash, oral ulcers, diffuse alopecia or edema in bilateral eyelids and lower limbs. Pertinent laboratory findings included an ESR of 101 mm/h. White blood cell count, platelet count, and hemoglobin were all normal. Parathyroid hormone (PTH) was 351 pg/ml. CaAfter admission, laboratory tests were positive for serum immunoglobulin G4 (IgG4), 38,500 mg/L , accounting for 66.9 % of total IgG. CT scanning showed there were enlarged lymph nodes in the left side of face, in bilateral orbital cavity, and bilateral temporal fossa. Scanning of CT indicated that there were thyroid nodules and enlarged lymph nodes in mediastinum, abdominal cavity, and retroperitoneum. The morphology of peripheral blood cell was normal.The patient accepted renal biopsy but refused lymph node biopsy. We obtained two renal biopsy specimens, and each specimen with length of about 1 cm contained 100 % cortex with 13 glomeruli. On light microscopy of hematoxylin-eosin staining, there was total 13 glomeruli, including 6 glomeruli completely fibrosis and 3 glomeruli peribulbar fibrosis. The mesangial cell and matrix showed no proliferation, there was no thickening of glomerular capillary. In Masson staining, no immune complexes deposited in capillary wall. In PASM staining, 60 % of tubules were atrophic, others showed dilatation with flat epithelial cells, renal interstitial showed diffuse fibrosis with a large number of plasma cell diffuse infiltration and IgG4 staining was positive (average 47 IgG4-positive plasma cells per high power field) Figs. 3.Fig. 1AThe result of electron microscopy showed: there was no significant proliferation and expansion in mesangial cells and matrix. The capillary wall basement membrane showed diffuse thickening and shrinkage, without significant electron dense deposit. Foot process of podocyte appeared diffuse effacement. Tubular epithelial cell and organelle swollen. Tubular basement membrane was almost normal. The interstitium showed focal edema, fibrosis with inflammatory cell infiltration.According to clinical manifestations, laboratory findings, and imaging results, the case was diagnosed as IgG4-related disease. He was managed with standard prednisolone with relative drugs. Ten days later, Scr and ESR decreased. Enlarged lymph nodes behind of left ear, right groin and left orbital cavity, became significantly smaller 10 days after starting steroid therapy, each was about 0.5 × 0.5 cm. Lower extremity numbness also mitigated.One month later, he returned for re-examination. Enlarged lymph nodes in the behind of left ear, right groin and left orbital cavity disappeared. Lower extremity numbness also disappeared. ESR decreased to 15 mm/h, Scr decreased to 311 µmol/L, urinary ACR was 0.2 g/gCr and level of IgG4 decreased to 8500 mg/L. Enlarged lymph nodes became smaller than before in CT. Two months later, level of IgG4 decreased to 2060 mg/L.IgG4-related disease is a disease associated with serum increased IgG4, which is involved in chronic and autoimmune disease in multiple organs and/or tissues ” in the year 2000. Because of the limited medical knowledge of IG4-related systemic disease in the year 2000, IgG4 staining did not completed at that time. Unfortunately the specimens at diagnosis were lost and we could not perform IgG4 staining. CT scanning at different timepoints showed enlarged lymph nodes in many organs. Laboratory tests was positive, 38,500 mg/L, for IgG4, >1350 mg/L, accounting for 66.9 % of total IgG (>40 %). In renal biopsy, 60 % tubules was atrophy, other tubules showed dilatation with flat epithelial cells, renal interstitial showed diffuse fibrosis with diffuse infiltration of a large number of plasma cell and IgG4 staining was positive Figs. 2. Previous reports showed that IgG4-related disease is a kind of benign lesions, but there has not been well-defined standard treatment so far. It was generally sensitive to glucocorticoids (Wallace and Stone It has been only 15 years since the first IgG4-related system diseases was reported Takahashi et al. , and theIgG4-related disease is a systemic fibro-inflammatory immune-mediated disorder and now recognized in many organs. It is characterised with multiple enlarged lymph nodes and well response to glucocorticoids. IgG4-related disease progresses slowly,and in the present patient the course of IgG4-related disease appears to be benign. The prognosis of IgG4-related disease is perhaps good as long as early detection and treatment.


Staphylococcus aureus (including MRSA) increasingly appears in individuals with no healthcare associated risks. Our study assessed risk factors for nasal carriage of resistant S. aureus in a multinational, healthy, community-based population, including ecological exposure to antibiotics.Antimicrobial resistance (AMR) has become a global public health concern which threatens the effective treatment of bacterial infections. Resistant S. aureus to antibiotic exposure and other risk factors (e.g. age and profession).Data were collected in eight European countries . Commensal AMR patterns were assessed by collecting 28,929 nasal swabs from healthy persons (aged 4+). Ecological exposure to antibiotics was operationalized as systemic antibiotic treatment patterns, extracted from electronic medical records of primary care practices in which the participants were listed (10–27 per country). A multilevel analysis related AMR in nasal commensal S. aureus isolates, 77% showed resistance to at least one antibiotic. 7.1% exhibited multidrug resistance (defined as resistance to 3 or more antibiotic classes), and we found 78 cases MRSA (1.3%). A large variation in antibiotic exposure was found between and within countries. Younger age and a higher proportion of penicillin prescriptions in a practice were associated with higher odds for carriage of a resistant S. aureus. Also, we found higher multidrug resistance rates in participants working in healthcare or nurseries.Of the 6,093 S. aureus, highlighting the need for cautious prescribing in primary care. Finally, since variation in AMR could partly be explained on a national level, policy initiatives to decrease AMR should be encouraged at the national level within Europe.This study indicates that in a population with no recent antibiotic use, the prescription behavior of the general practitioner affects the odds for carriage of a resistant  Staphylococcus aureus  was confined to hospitals and long-term-care facilities. In the 21th century however, MRSA infections have also appeared in community-dwelling individuals with no healthcare associated risks such as a recent hospitalization : t: tS. aurS. aureus was isolated. Excluding participants without AMR data  resulted in a total number of 6,093 individuals in our study. This sample consisted of 5,224  adults (aged 18+) and 839  children (aged 4 to 17), and 30 persons  whose age was unknown. 51% of all participants was female.Data were obtained from swabs from a total number of 28,929 persons: in twenty-one percent  a S. aureus, and categorized each microbe in two different ways:We assessed nasal carriage of Resistance to none of the tested antibiotics versus resistance to at least one the tested antibiotics.Multidrug resistance:Resistance to 0, 1 or 2 antibiotic classes versus resistance to 3 or more antibiotic classes (ATC 3 level).Whether the isolated microbe is resistant at all:S. aureus to be dynamic and occurring on multiple bodily sites. While the prevalence might be underestimated by using nasal swabs [S. aureus to be representative of all carriage.We included multidrug resistance as a variable instead of MRSA, since in our study only 78 MRSA isolates were found. They have been described in more detail elsewhere . Previoual swabs , we assuEcological antibiotic exposure was operationalized as antibiotic treatment volume and pattern of the primary care practices in which the participant was listed. We assume that the number of antibiotic prescriptions per practice is an approximation of exposure for the participants from that practice. Data about systemic antibiotic treatments (ATC codes J01) were extracted from the electronic medical records of the participating primary care practices from which this was possible (90%). Given that AMR developed after exposure to an antibiotic can linger up to one year , we inclThe load of antibiotics is defined as the total number of packages/100 active patients prescribed by practice in 2010. Active patients are defined as those who have visited the practice at least once during the year 2010.S. aureus in the community [Secondly, we calculated the percentage of these prescribed packages which consists of beta-lactamase susceptible penicillin (since this is known to be the antibiotic with the highest resistance levels in ommunity ).To assess the association between antibiotic exposure and AMR we operationalized this exposure in two different ways:S. aureus, and we also included gender and the number of GP visits in the last year, since in all participating countries a prescription is needed for an antibiotic. We found no previous evidence for variables on practice level to be correlated to carriage of a resistant S. aureus.All participants completed a short questionnaire with background information. Based on previous literature, we selected several variables which were used in our model. On the patient level, several variables have been demonstrated to increase the odds of AMR: a younger age , living The dependent variable (AMR) was nested on three levels: person, primary care practice, and country. A multilevel logit analysis was conducted (using MLwiN version 2.33), relating AMR to ecological antibiotic exposure, controlling for other risk factors. Both analyses were based on a dichotomous dependent variable: for model 1, it was coded 0 if an isolate shows no resistance and 1 if resistance to at least 1 antibiotic is found. For model 2, the response variable was coded 1 if an isolate is demonstrated to be multidrug resistant (otherwise coded 0). For useful interpretation of the results, the continuous variables in the analysis  were transformed into quartile scores see .S. aureus in the community: a resistance rate of <20% to all antibiotics was found, except for (beta-lactamase susceptible) penicillin, to which 78% resistance was demonstrated  see ). AntibiA large variation between countries was found: practices in Sweden and France stood out with a low number (<20) of prescribed packages per 100 active patients, whilst practices in Croatia, Hungary and Spain on average prescribed over 60 packages for every 100 active patients. Finally, the extent to which GPs opted for penicillin to treat bacterial infections differed: from 1 in every 5 prescribed packages in Croatia, to almost half of the prescriptions in Sweden. At the same time however, Sweden displayed the lowest antibiotic load, and the lowest AMR levels of all countries. Spain on the other hand demonstrated high AMR levels and a high average exposure to antibiotics .Descriptive statistics of all variables are shown in S. aureus resistant to at least one antibiotic. The results showed that participants with a younger age had a higher risk to carry a resistant S. aureus. Next to this, persons listed in practices with higher proportional penicillin exposure had a higher risk to carry a resistant S. aureus. Both results show relatively small odds, and are based on quartile scores for which a linear relationship was assumed. Overall, in the first model, almost no variation was found on the practice level, but a considerable variation was seen at the country level.The first model assessed risk factors for carriage of a S. aureus. On the other hand, although penicillin prescriptions were related to AMR in general (model 1), the level of exposure to antibiotics had no significant effect on colonization of multidrug resistant bacteria.Results concerning multidrug resistance (Model 2) showed that certain professions are associated with multidrug resistance: participants working in health care or nurseries had a 1.7–1.9 times higher risk for carrying a multidrug resistant S. aureus in a community population with no health-care associated risks. We identified several risk factors associated with carriage of a resistant S. aureus: age, profession, treatment with penicillin and country.Our study integrates and expands current knowledge by investigating the nasal commensal microbiota, which are a reservoir for resistant pathogens ,9, takinOur results show that resistance to any antibiotic is associated with lower age. Previous studies support this result . We founRegarding multidrug resistance, we found that working in health care or a nursery is associated with multidrug resistance. Finally, most variance in resistance is found at the country level. This implies that although AMR is associated with antibiotic exposure in the GP practice, commensal AMR can be considered a national phenomenon. The preferred level of action should therefore be national : alreadyS. aureus, and the broad perspective by including data from 8 European countries and focussing on a population without health-care associated risks. Within these countries, extensive prescription data and AMR data were collected, enabling unique analyses.A strength of this study is the integration of knowledge regarding community-associated S. aureus is dynamic [S. aureus. We assume that AMR patterns in the nasal commensal flora are comparable with those of pathogenic S. aureus. However, for empirical treatment purposes, AMR data on pathogenic isolates is also needed. Furthermore, we did not use antibiotic treatment data on patient level and used practice level antibiotic prescription data as an approximation of ecological exposure to antibiotics at the individual level. The healthcare system of the country could affect the representativeness of our measure of exposure to antibiotics. In some countries GPs function as a gatekeeper to the healthcare system, while elsewhere other medical disciplines also prescribe antibiotics. Although over the counter sales of antibiotics without prescriptions is prohibited, we cannot rule out the possibility of our patients being exposed to antibiotics elsewhere. Studies indicating major effects from other sources are lacking, therefore we assume that our measurement is a good approximation of the ecological exposure to antibiotics in the community, with a possible underestimation.The study is limited regarding several assumptions that have been made in our analysis: although carriage of  dynamic , we assuOur data collection of antibiotic prescriptions showed the difficulties of establishing a congruent database given the different ways of collecting treatment data across Europe (data was extracted from different electronic systems based on different classification models). Not all participating practices were able to deliver complete treatment data, which could indicate a selection bias towards more involved GPs. We assume the participating practices to be representative for the whole country regarding prescription behaviour. A comparison with ESAC reports  supportsS. aureus; this should create a higher awareness under general practitioners. AMR can also spread to patients who are not directly exposed: in this ecological dilemma the benefits for the patient have to be weighed against the drawbacks for the total population. After correcting for other risk factors, we found mainly national variation in resistant S. aureus, indicating that national actions are recommended.Our results demonstrated several risk factors for AMR in nasal commensal S1 Fig(DOCX)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file.S4 Table(DOCX)Click here for additional data file.