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Calcium oxalate (CaOx) is the most common component of kidney stones. Oxidative stress, inflammation and autophagy-induced cell death are the major causes of CaOx crystal deposition and CaOx crystal deposition can further lead to kidney injury. Trimethylamine N-oxide (TMAO), a gut microbiota-derived metabolite, plays an important role in the pathogenesis of many diseases, such as atherosclerosis, diabetes and chronic kidney disease, but the effect of TMAO on hyperoxaluria-induced CaOx crystal deposition and kidney injury remains unknown. We hypothesize that TMAO aggravates CaOx crystal deposition via promoting CaOx-mediated cell death. C57Bl/6 mice were given high-oxalate diet as a model of hyperoxaluria. TMAO was provided via drinking water. Serum TMAO levels increased 15 days after CaOx treatment (6.30 ± 0.17 μmol/L vs. 34.65 ± 8.95 μmol/L). High-oxalate diet induced inflammation, CaOx deposition and kidney injury, which TMAO aggravated. In accordance, TMAO intensified high-oxalate diet induced oxidative stress, autophagy and apoptosis. Moreover, TMAO enhanced CaOx crystal adhesion to HK-2 cells and reduced cell viability (from 88.9 ± 1.6% to 75.0 ± 2.7%). Protein kinase R-like endoplasmic reticulum kinase (PERK) may mediate these TMAO effects, as TMAO promoted PERK phosphorylation. Consistently, PERK knockdown alleviated TMAO-evoked CaOx-autophagy, apoptosis and oxidative stress in HK-2 cells. In conclusion, TMAO can aggravate hyperoxaluria-induced kidney injury by triggering the PERK/ROS pathway, which enhances autophagy, apoptosis and inflammation, and facilitates CaOx crystal deposition in renal tubular cells.

Background: To provide tacrolimus is first-line treatment after liver and kidney transplantation. However, hypertension and nephrotoxicity are common tacrolimus side effects that limit its use. Although tacrolimus-related hypertension is well known, the underlying mechanisms are not. Here, we test whether tacrolimus-induced hypertension involves the RhoA (Ras homolog family member A)/ROCK (Rho-associated protein kinase) pathway in male C57Bl/6 mice. methods: Intra-arterial blood pressure was measured under anesthesia. The reactivity of renal afferent arterioles and mesenteric arteries were assessed in vitro using microperfusion and wire myography, respectively. Results: Tacrolimus induced a transient rise in systolic arterial pressure that was blocked by the RhoA/ROCK inhibitor Fasudil (12.0±0.9 versus 3.2±0.7; P P 2+ mobilization and sensitization. Fasudil prevented increased Ang II-sensitivity and blocked Ca 2+ mobilization and sensitization. Preincubation of mouse aortic vascular smooth muscle cells with tacrolimus activated the RhoA/ROCK/MYPT-1 (myosin phosphatase targeting subunit 1) pathway. Further, tacrolimus increased cytoplasmic reactive oxygen species generation in afferent arterioles (107±5.9 versus 163±6.4; P P Conclusions: The RhoA/ROCK pathway may play an important role in tacrolimus-induced hypertension by enhancing Ang II-specific vasoconstriction, and reactive oxygen species may participate in this process by activating the RhoA/ROCK pathway.

Gut microbiota produce Trimethylamine N-oxide (TMAO) by metabolizing dietary phosphatidylcholine, choline, l-carnitine and betaine. TMAO is implicated in the pathogenesis of chronic kidney disease (CKD), diabetes, obesity and atherosclerosis. We test, whether TMAO augments angiotensin II (Ang II)-induced vasoconstriction and hence promotes Ang II-induced hypertension. Plasma TMAO levels were indeed elevated in hypertensive patients, thus the potential pathways by which TMAO mediates these effects were explored. Ang II (400 ng/kg−1min−1) was chronically infused for 14 days via osmotic minipumps in C57Bl/6 mice. TMAO (1%) or antibiotics were given via drinking water. Vasoconstriction of renal afferent arterioles and mesenteric arteries were assessed by microperfusion and wire myograph, respectively. In Ang II-induced hypertensive mice, TMAO elevated systolic blood pressure and caused vasoconstriction, which was alleviated by antibiotics. TMAO enhanced the Ang II-induced acute pressor responses (12.2 ± 1.9 versus 20.6 ± 1.4 mmHg; P, Graphical abstract Image 1, Highlights • Orally administered TMAO aggravates Ang II-induced hypertension. Antibiotics alleviate Ang II-induced hypertension by reducing TMAO generation. • High concentrations of TMAO constrict afferent arterioles and mesenteric arteries and increase blood pressure. • Low concentrations of TMAO enhance Ang II-induced vasoconstriction and acute pressor response via activating PERK/ROS/CaMKII/PLCβ3/Ca2+ pathway.

Aims Reduced A Disintegrin And Metalloproteinase with a ThromboSpondin type 1 motif member 13 (ADAMTS13) levels are observed in kidney disease. We test whether recombinant human ADAMTS13 (rhADAMTS13) mitigates renal injury in chronic kidney disease (CKD) and the potential mechanisms. Methods CKD was established 3 months after ischaemia/reperfusion (IR). ADAMTS13 and von Willebrand factor (vWF) levels, renal function and morphological changes were analysed. Afferent arteriolar responses to angiotensin II (Ang II) and acetylcholine (ACh) were measured. Oxidative stress-related molecules were detected. Results Higher vWF and lower ADAMTS13 levels were observed in CKD mice, which were markedly attenuated by rhADAMTS13. rhADAMTS13 alleviated renal dysfunction, as documented by decreased blood urea nitrogen (BUN), serum creatinine, kidney injury molecule-1 (KIM-1) and neutrophil gelatinase-associated lipocalin (NGAL) levels in CKD mice. Moreover, rhADAMTS13 attenuated transforming growth factor (TGF)-β1/Smad3 activation. Plasma vWF: ADAMTS13 ratio showed positive correlations with malondialdehyde (MDA), hydrogen peroxide (H2 O2 ) and proteinuria, and correlated inversely with superoxide dismutase (SOD) and catalase (CAT). Finally, rhADAMTS13 inhibited reactive oxygen species (ROS) levels and improved microvascular functional disorders, accompanied by the inhibition of glycogen synthase kinase (GSK) 3β hyperactivity and upregulation of nuclear factor erythroid 2-related factor 2 (Nrf2) expression. Conclusions Acute kidney injury (AKI) reduces the expression of ADAMTS13 that contributes to progressive CKD, microvascular dysfunction, oxidative stress, inhibition of Nrf2 activity and renal histopathological damage. All of which can be alleviated by administration of rhADAMTS13.

Aims In patients with renal disease, high serum phosphate shows a relationship with cardiovascular risk. We speculate that high phosphate (HP) impairs arterial vasodilation via the endothelium and explore potential underlying mechanisms. Methods Isolated vessel relaxation, endothelial function, glomerular filtration rate (GFR), oxidative stress status and protein expression were assessed in HP diet mice. Mitochondrial function and protein expression were assessed in HP-treated human umbilical vein endothelial cells (HUVECs). Results High phosphate (1.3%) diet for 12 weeks impaired endothelium-dependent relaxation in mesenteric arteries, kidney interlobar arteries and afferent arterioles; reduced GFR and the blood pressure responses to acute administration of acetylcholine. The PPARα/LKB1/AMPK/eNOS pathway was attenuated in the endothelium of mesenteric arteries from HP diet mice. The observed vasodilatory impairment of mesenteric arteries was ameliorated by PPARα agonist WY-14643. The phosphate transporter PiT-1 knockdown prevented HP-mediated suppression of eNOS activity by impeding phosphorus influx in HUVECs. Endothelium cytoplasmic and mitochondrial reactive oxygen species (ROS) were increased in HP diet mice. Moreover HP decreased the expression of mitochondrial-related antioxidant genes. Finally, mitochondrial membrane potential and PGC-1α expression were reduced by HP treatment in HUVECs, which was partly restored by AMPKα agonist. Conclusions HP impairs endothelial function by reducing NO bioavailability via decreasing eNOS activity and increasing mitochondrial ROS, in which the AMPK-related signalling pathways may play a key role.

The natural course of radiation encephalopathy following nasopharyngeal carcinoma (NPC) radiotherapy remains poorly understood. The present study aimed to investigate the magnetic resonance imaging (MRI) characteristics and evolution of radiation encephalopathy. A series of 162 follow-up MRI examinations from 68 NPC patients with radiation encephalopathy in the temporal lobes were analyzed retrospectively. Each component of radiation encephalopathy was defined as follows: i) contrast enhanced lesions were enhanced lesions on contrast enhanced T1-weighted images (T1WI); ii) white matter lesions were lesions of homogeneous hyperintensity on T2-weighted images (T2WI) and hypointensity on T1WI; iii) cysts were round or oval well-defined lesions of hyperintensity on T2WI; iv) hemosiderin deposition were nodular or annular hypointense lesions with lower hypointense than normal white matter on both T1WI and T2WI; v) gray matter lesions were defined as disruption or erosion of hyperintensity in the cortex on T2WI. Contrast enhanced lesions, white matter lesions, gray matter lesions, cysts and hemosiderin deposition were detected in 105 (100.0%), 98 (93.3%), 94 (89.5%), 2 (1.7%) and 2 (1.7%) cases of the 105 initial diagnosed temporal lobe lesions. Contrast enhanced lesions were the most commonly observed, followed by white matter lesions, gray matter lesions, temporal lobe atrophy, cysts and hemosiderin deposition. In addition, 12 new lesions were identified during the follow-up, 4 of which presented as solid enhanced nodular lesions. Importantly, in 11 of the 117 (9.4%) affected temporal lobes, solid enhanced nodular lesions were observed to be the only initial abnormalities to occur. For those enhanced nodular lesions measuring 2.0 cm exhibited a necrotic core. To the best of our knowledge, the present study revealed for the first time solid enhanced nodular lesions as the earliest MRI abnormalities of radiation encephalopathy following NPC radiotherapy.

In the era of intensity-modulated radiotherapy, distant metastasis is currently the main cause of treatment failure for nasopharyngeal carcinoma (NPC). Additional therapeutic strategies are required to control the metastasis and improve survival. One strategy is targeted therapy, for example against c-Kit. In the current study, the frequency of c-Kit expression was determined immunohistochemically in 106 NPC patients. c-Kit expression changes during the course of radiation therapy were detected in 41 cases via weekly biopsy. Twelve cases (11.3%) had c-Kit expression scores of 3+ and 16 (15.1%) had scores of 2+. Thus, c-Kit overexpression (2+ or 3+) was observed in 28 (26.4%) patients. There were 35 (33.0%) and 43 (40.6%) patients with c-Kit expression scores of 1+ and 0, respectively. Furthermore, a trend of decreased c-Kit expression was observed after commencing radiotherapy according to the 41 NPC patients who were biopsied weekly. Therefore, c-Kit overexpression was identified to be common in NPC, and evaluating c-Kit as a therapeutic target for metastatic NPC via c-Kit overexpression subsequent to first line treatment may be of interest. To the best of our knowledge, the present study is the first to demonstrate a trend of decreased c-Kit expression during the course of radiotherapy.

Background There is currently no worldwide consensus for the management of hepatocellular carcinoma (HCC) with portal vein tumor thrombus (PVTT). We evaluated the efficacy of stereotactic body radiotherapy (SBRT) as the initial treatment for HCC with extensive PVTT based on a relatively large number of patients. Methods In our multidisciplinary approach for patients with hepatobiliary tumors, SBRT is recommended for unresectable HCC with PVTT or those with contraindication for transarterial chemoembolization (TACE). The aim is to shrink the tumor thrombus and preserve adequate portal venous flow, thus facilitating subsequent treatments such as TACE and tumor resection. In the present study, 70 continuous cases of HCC patients with extensive PVTT initially treated with SBRT were studied. The median follow-up period was 9.5 months (range, 1.0–21.0 months). The dynamic changes of tumor thrombosis with time after SBRT were also analyzed. Results The median survival time for the whole group was 10.0 months (95% CI, 7.7–12.3 months), with a 6- and 12-month overall survival (OS) rate of 67.3%, and 40.0% respectively. Patients who received combined SBRT and TACE showed significantly longer OS than those without indication for TACE after SBRT (12.0 ± 1.6 vs. 3.0 ± 1.0 months). Patients with good response to radiation usually had better survival. SBRT was well tolerated in our patient series. Conclusions In conclusion, SBRT used as the initial treatment for HCC patients with extensive PVTT originally unsuitable for resection or TACE can achieve adequate thrombus shrinkage and portal vein flow restoration in the majority of cases. It could thus offer the patients an opportunity to undergo further treatment such as resection or TACE procedure. Such therapeutic strategy may result in survival advantage, especially for those who do receive combined modality with SBRT.

Background: The differences in genetic characteristics between primary and recurrent gliomas have been poorly characterized. We investigated whether genetic characteristics in recurrent gliomas would differ from those of primary gliomas, as a means of identifying unique markers in recurrent glioma to permit a better understanding of disease progression and therapeutic considerations. Methods: Multiple genes involved in eight important pathways in primary and their corresponding paired recurrent glioma samples from 42 patients were analyzed by immunohistochemical staining. Findings: In recurrent glioma, we saw a significantly increased expression of PD-1, PD-L1, VEGF, VEGFR2, CD133, MGMT, and Rad51 when compared with their corresponding primary tumors. VEGF expression was correlated with VEGFR2 expression in both primary and recurrent gliomas, and their co-expression was associated with shorter progression-free survival (PFS) and overall survival (OS). PD-1 expression was correlated with PD-L1 expression in recurrent gliomas. As compared to primary gliomas, the overall expression of MGMT was increased in the adjuvant therapy group but was not statistically different from the no-adjuvant therapy group. The group in which MGMT increased had a shorter PFS and OS than did the group in which MGMT was not increased. Co-expression of P53 and Ki-67 in recurrent gliomas was associated with poor median OS after second resection. Interpretation: The gene expression characteristics of recurrent gliomas were significantly different from those of their corresponding primary gliomas, a result that has important implications for patient survival. Treatment based on these newly identified changes may offer better therapeutic options for recurrent glioma. Funding Statement: This work was surported by the National Natural Science Foundation of China (81572952). Declaration of Interests: The authors declare that they have no conflict of interest. Ethics Approval Statement: This study was approved by the Ethics Committee of SAHZU, and was carried out in accordance with the Declaration of Helsinki.

Objectives To explore the areas at highest risk for postoperative pancreatic cancer local recurrence according to the spatial location of local failures, with the aim to provide a precise target volume for pancreatic cancer adjuvant radiotherapy. Methods Patients with pancreatic cancer who had undergone surgery for the primary tumor in pancreas at our institution from January 2010 to August 2015 were retrospectively analyzed. All local recurrences were plotted on the computed tomography (CT) image of a representative patient according to their relative coordinates to superior mesenteric artery (SMA) or celiac axis (CA). Adjuvant radiation clinical target volume (CTV)-90 and CTV-80 were created to cover 90 % and 80 % plotted recurrences. This planning approach was applied in four simulated cases with comparison to the plan according to RTOG 0848 contouring consensus guidelines. Raystation v4.5.1.14 was used for analyzing high throughput physics data. Results Eighty-three patients with local recurrence were included from 305 postoperative pancreatic cancer patients who did not receive adjuvant radiotherapy. Thirty-one (37 %) patients did not have adjuvant therapy at all, 52 (63 %) patients undergone adjuvant chemotherapy alone. Spatial location of local failure was created. Most recurrences occurred near CA or SMA. CTV-90 was generated through expanding the combined SMA and CA contours by 30 mm right-lateral, 21 mm left-lateral, 20 mm anterior, 13 mm posterior, 10 mm superior, and 20 mm inferior. CTV-80, smaller in volume, was also created for simultaneous integrated boost. Through comparison and analysis of the simulated cases, the radiation volumes proposed were much smaller than those with RTOG 0848 contouring consensus guidelines (average volume: PTV-80 = 120 ml, PTV-90 = 220 ml, RTOG PTV = 490 ml). Accordingly, the organs at risk received less irradiation dose with the proposed CTV-90 and CTV-80. Conclusions Smaller adjuvant radiotherapy CTVs targeting the high-risk local failure areas of postoperative pancreatic cancer were proposed, according to the three-dimensional spatial location of local recurrences. This may help to minimize radiation-related toxicities, achieve dose escalation, and finally reduce local recurrence.