Distinct histopathological phenotypes of severe alcoholic hepatitis suggest different mechanisms driving liver injury and failure

Intrahepatic neutrophil infiltration has been implicated in severe alcoholic hepatitis (SAH) pathogenesis; however, the mechanism underlying neutrophil-induced injury in SAH remains obscure. This translational study aims to describe the patterns of intrahepatic neutrophil infiltration and its involvement in SAH pathogenesis. Immunohistochemistry analyses of explanted livers identified two SAH phenotypes despite a similar clinical presentation, one with high intrahepatic neutrophils (Neu^hi), but low levels of CD8⁺ T cells, and vice versa. RNA-Seq analyses demonstrated that neutrophil cytosolic factor 1 (NCF1), a key factor in controlling neutrophilic ROS production, was upregulated and correlated with hepatic inflammation and disease progression. To study specifically the mechanisms related to Neu^hi in AH patients and liver injury, we used the mouse model of chronic-plus-binge ethanol feeding and found that myeloid-specific deletion of the Ncf1 gene abolished ethanol-induced hepatic inflammation and steatosis. RNA-Seq analysis and the data from experimental models revealed that neutrophilic NCF1-dependent ROS promoted alcoholic hepatitis (AH) by inhibiting AMP-activated protein kinase (a key regulator of lipid metabolism) and microRNA-223 (a key antiinflammatory and antifibrotic microRNA). In conclusion, two distinct histopathological phenotypes based on liver immune phenotyping are observed in SAH patients, suggesting a separate mechanism driving liver injury and/or failure in these patients.     

Connectomic assessment of injury burden and longitudinal structural network alterations in moderate‐to‐severe traumatic brain injury

Traumatic brain injury (TBI) is a major public health problem. Caused by external mechanical forces, a major characteristic of TBI is the shearing of axons across the white matter, which causes structural connectivity disruptions between brain regions. This diffuse injury leads to cognitive deficits, frequently requiring rehabilitation. Heterogeneity is another characteristic of TBI as severity and cognitive sequelae of the disease have a wide variation across patients, posing a big challenge for treatment. Thus, measures assessing network‐wide structural connectivity disruptions in TBI are necessary to quantify injury burden of individuals, which would help in achieving personalized treatment, patient monitoring, and rehabilitation planning. Despite TBI being a disconnectivity syndrome, connectomic assessment of structural disconnectivity has been relatively limited. In this study, we propose a novel connectomic measure that we call network normality score (NNS) to capture the integrity of structural connectivity in TBI patients by leveraging two major characteristics of the disease: diffuseness of axonal injury and heterogeneity of the disease. Over a longitudinal cohort of moderate‐to‐severe TBI patients, we demonstrate that structural network topology of patients is more heterogeneous and significantly different than that of healthy controls at 3 months postinjury, where dissimilarity further increases up to 12 months. We also show that NNS captures injury burden as quantified by posttraumatic amnesia and that alterations in the structural brain network is not related to cognitive recovery. Finally, we compare NNS to major graph theory measures used in TBI literature and demonstrate the superiority of NNS in characterizing the disease.     

Phase II study of preoperative radiotherapy and concomitant weekly intravenous oxaliplatin combined with oral capecitabine for stages II–III rectal cancer

Introduction

A prospective phase II study was conducted to assess the clinical activity and tolerability of oxaliplatin, capecitabine, and radiotherapy (RT) for neoadjuvant therapy of stages II?CIII rectal cancer.

Materials and methods

Patients with histologically confirmed stages II?CIII (T3?CT4 and/or N+) resectable rectal adenocarcinoma were eligible. Capecitabine was administered at 825?mg/m² twice daily for 5?days/week and oxaliplatin at 50?mg/m² on day 1 weekly for 5?weeks starting the first day of RT (before RT). RT consisted of a total dose of 45?Gy delivered in 25 fractions of 1.8?Gy, 5?days per week, for 5?weeks.

Results

A total of 46 patients were included (35 male, 10 female, median age 62?years). TNM Stage was T3 in 43 patients and T4 in 2. Twenty-eight patients had suspected nodal involvement. The intended chemoradiation treatment was completed in 94?% patients. Grade 3/4 toxicity included lymphocytopenia (6 patients), diarrhea (4 patients), emesis (2 patients), asthenia (3 patients), anorexia (1 patient), and hepatic toxicity (1 patient). Grade 1 neurotoxicity occurred in 18 patients, Grade 2 neurotoxicity in 3, and Grade 1 palmoplantar erythrodysesthesia in 2. Forty-two patients underwent surgery (complete resection 95?%, sphincter-saving operation 55?%). The overall pathologic response rate was 83?%, with a pathologic complete response (pCR) rate of 11.9?% (95?% CI 4.0?C25.6).

Conclusions

The pCR rate observed with oxaliplatin plus capecitabine and RT did not reach the pre-specified criteria of efficacy in this trial, which is in line with recent results of randomized phase III trials.     

Mechanism of voltage sensing in Ca2+- and voltage-activated K+ (BK) channels

In neurosecretion, allosteric communication between voltage sensors and Ca²⁺ binding in BK channels is crucially involved in damping excitatory stimuli. Nevertheless, the voltage-sensing mechanism of BK channels is still under debate. Here, based on gating current measurements, we demonstrate that two arginines in the transmembrane segment S4 (R210 and R213) function as the BK gating charges. Significantly, the energy landscape of the gating particles is electrostatically tuned by a network of salt bridges contained in the voltage sensor domain (VSD). Molecular dynamics simulations and proton transport experiments in the hyperpolarization-activated R210H mutant suggest that the electric field drops off within a narrow septum whose boundaries are defined by the gating charges. Unlike Kv channels, the charge movement in BK appears to be limited to a small displacement of the guanidinium moieties of R210 and R213, without significant movement of the S4.

Excitable tissues accomplish their signaling functions thanks in part to the interplay of several voltage-sensitive ion channels (1–6). Hence, to understand these processes, it is crucial to establish how voltage-sensitive ion channels sense changes in the electric field across the membrane, an issue that has been a matter of extensive study and intense debate for decades. The most widely accepted mechanism proposes the existence of voltage-sensor domains (VSDs), modules that undergo two or more discrete conformational states in response to changes in the membrane voltage. The simplest model considers two states: active (A), which promotes pore opening, and resting (R), which promotes channel closing. To accomplish its function, VSDs contain voltage-sensitive particles, which move in response to changes in the electric field. This movement triggers the interconversion between the two discrete conformational states. These voltage-sensing particles are typically the guanidine groups of arginine residues within the S4 transmembrane segment, which undergo a combination of rotational, translational, and tilting movement in response to changes in membrane voltage (7–14).The large-conductance Ca²⁺- and voltage-activated K⁺ (BK) channels have a wide distribution in mammalian tissues (15–18), where they participate in a diversity of physiological processes. Their malfunction is often related to diverse pathological conditions (19, 20). BK channel open probability is independently regulated by membrane depolarization and intracellular Ca²⁺ concentration (21, 22), each stimulus being detected by specialized modules. Like other voltage-sensitive K⁺ (Kv) channels, BK is an homotetramer in which each of its α subunits consists of a pore domain (PD; S5-S6 transmembrane segments), a voltage-sensing domain (VSD; S1–S4 transmembrane segments) containing a positively charged S4, and a cytosolic C-terminal regulatory domain, which contains the Ca²⁺-binding sites (23, 24). Also, like some members of other K⁺ channel families (25, 26), the VSD and PD of BK are non–domain swapped (23, 24). BK channels display some distinctive structural and functional features: Despite sharing the selectivity filter sequence with Kv channels, BK unitary conductance and selectivity are exquisitely high (27–30). The BK α subunit has an additional transmembrane segment S0 [therefore, its N terminus faces the extracellular medium (31)], and the voltage sensitivity in BK channels is significantly lower than that of Kv channels, presumably because of their lower number of gating charges (32).Although thoroughly studied, research into BK VSD and its voltage dependence has faced several technical obstacles. The relatively small gating charge per channel (32) and the large conductance of the BK pore makes isolating of the gating currents from the ionic currents a tough experimental challenge. In addition, because mutations of VSD residues can produce very large shifts in both the gating charge-voltage (Q(V)) and the conductance-voltage G(V)) relationships (33), it is necessary to use extreme voltages to accurately measure the voltage dependence of some mutants. Consequently, the identification of BK gating charges has been addressed by using indirect approaches (33, 34). The combination of electrophysiology measurements and kinetic modeling suggests a decentralized VSD in the BK channel, where four charged residues (D153 and R167 in S2, D186 in S3, and R213 in S4) act as voltage sensor particles (33). A recent report of the atomistic cryo-electron microscopy (cryo-EM) structures of the human BK channel and its homolog in Aplysia californica (AcSlo) revealed minor structural differences between the VSD in both the Ca²⁺-bound (open pore) and the Ca²⁺-unbound (closed pore) conformations (23, 24, 35). This result can be explained if the conformational changes of the BK VSD upon activation are small compared to those that occur during the activation of other channels, such as HCN channels (12–14).In this study, we identified voltage-sensing particles in the BK channel by using a direct functional approach, involving gating of current measurements and analysis of the Q(V) curves spanning 800 mV in the voltage axis. Systematic neutralization of the individual charged residues in the VSD (S1–S4) revealed that only the neutralization of two arginines in S4 (R210 and R213) changed the voltage dependence of the Q(V) curves. Neutralization of other VSD charges point to roles in tuning of the half-activation voltage of the VSD and its allosteric coupling with the PD. Molecular dynamics (MD) simulations based on the cryo-EM structures of the human BK channel (35) as templates suggested that R210 and R213 lie in a very narrow septum separating intra- and extracellular water-filled vestibules. This interpretation is consistent with the robust hyperpolarization-activated proton currents generated when R210 is mutated to the protonable amino acid histidine. Overall, our results point to a unique and distinctive mode of activation in BK: In contrast to Kv channels, where positive charges move one by one through a charge transfer center (absent in BK channels) that spans the entire electric field (36, 37), charge movement in BK channels is limited to the small displacement of R210 and R213, which itself constitutes a narrow septum where the electric field drops.     

Factors associated with satisfaction with care among patients with epilepsy

OBJECTIVE: The purpose of the work described here was to determine those variables associated with satisfaction with care among patients with epilepsy. METHODS: We interviewed patients followed at a tertiary epilepsy center. Predictor variables included age, gender, race, education, income, insurance, seizure frequency, and Quality of Life in Epilepsy-10 inventory (QOLIE-10) results. Target variables were the subscales of the Short Form Patient Satisfaction Questionnaire (PSQ-18). We used univariate analysis to identify those variables significantly associated with the subscales and multiple linear regression to determine those independently significant. RESULTS: The study population comprised 193 patients. Lower education and better QOLIE-10 scores were independently associated with general satisfaction with care. The mental health scale was associated with general satisfaction with care. Lower educational level was the only variable independently associated with patient satisfaction with communication, the financial aspect of care, and time spent with physician. CONCLUSION: Lower educational level and better quality of life are the main variables associated with higher general satisfaction with care among patients with epilepsy.     

Induction of Apoptosis in HT-29 Cells by Extracts from Isothiocyanates-rich Varieties of Brassica Oleracea

Among the vegetables with anti-carcinogenic properties, members of the genus Brassica are the most effective at reducing the risk of cancer. This property may be explained by their principle bioactive compounds, isothiocyanates (ITCs). The aim of this study was to measure the amounts of ITCs in extracts from vegetables of the Brasssica genus and assay them for potency of induction of apoptosis in a colorectal cancer cell line (HT-29). ITCs were determined by the cyclocondensation assay with 1,2-benzenedithiol and induction of apoptosis by assessment of cell viability, caspase-3 activity and DNA fragmentation. Purple cabbage extract showed the highest ITC concentration per gram, fresh weight, followed by black cabbage and Romanesco cauliflower. At ITC concentrations of 7.08 μ g/mL these extracts decreased cell viability and induced caspase-3 and DNA fragmentation at 48h. Brussels sprouts showed the strongest effects on cell viability and caspase-3 activity. Varieties of Brassica Oleracea are rich sources of ITCs that potently inhibit the growth of colon cancer cells by inducting apoptosis. All the extracts showed anticancer activity at ITC concentrations of between 3.54 to 7.08 μ g/mL, which are achievable in vivo. Our results showed that ITC concentration and the chemopreventive responses of plant extracts vary among the varieties of Brassica Oleracea studied and among their cultivars.     

p-Amminobenzoic Acid as a Protective Agent in Ozone Toxicity

The ability of p-aminobenzoìc acid (PABA) to profeet against ozone toxicity was demonstrated in two experimental systems p-Aminobenzoic acid, and the related compounds p-aminohippuric acid and anthranïlïc acid, inhibited the ozone-induced loss of activity of the enzyme acetylcholinesterase in human erythrocytes exposed in vitro to 40 ppm of ozone. Protection in vivo was demonstrated In a study in which 15 rats injected intraperitoneally with 2 ml of 20 mM PABA prior to ozone exposure died in a mean of 426 minutes as compared to a mean of 261 minutes for 15 control animals similarly exposed to 15 ppm of ozone. These findings provide additional presumptive evidence of the importance of free radicals in the mechanism of ozone toxicity. They further suggest that PABA and related compounds warrant exploration as possible protective agents in susceptible populations exposed to significant photochemical air pollution.