Follow-up of women with cervical adenocarcinoma in situ treated by conization: A single centre clinical experience.

OBJECTIVE: Hysterectomy has been the historical gold standard final step in the treatment algorithm of adenocarcinoma in situ (AIS) recommended by most North American colposcopy guidelines. AIS disproportionately affects young childbearing age women, therefore a fertility sparing treatment option is desirable. Our study examines the impact of conservative treatment of AIS with conization followed by serial surveillance. METHODS: A retrospective chart review was completed of patients treated for AIS from 2006 to 2020. Charts were identified by pathologic diagnosis of AIS on cervical and uterine specimens. Charts were excluded if AIS was not treated with conization, if AIS was not confirmed on initial conization specimen, or if invasive disease was found at initial conization. RESULTS: 121 patient charts were analyzed. Median age of patients at first conization and hysterectomy was 34.8 and 40.9, respectively. First conization was by Cold Knife Cone in 58% of patients, and by Loop Electrosurgical Excisional Procedure in 42% of patients. Median follow-up period in our study was 609 days. 5% of patients had recurrence, with only one patient who recurred as cancer. One case of recurrence had a positive initial conization margin. Median time to recurrence was 700 days. 47% of patients underwent eventual hysterectomy. Residual AIS was found in 23% of hysterectomy specimens. Adenocarcinoma was diagnosed on hysterectomy specimen in four patients. CONCLUSION: Our study demonstrates the oncologic safety of treating AIS with conization and serial surveillance. Routine hysterectomy completed as a part of the AIS treatment algorithm, as in current clinical guidelines, is unnecessary.

Demonstration of chemotherapeutic mediated lymphatic changes in meningeal lymphatics in vitro, ex vivo, and in vivo.

Systemic chemotherapeutics target cancer cells but are also known to impact other cells away from the tumor. Questions remain whether systemic chemotherapy crosses the blood-brain barrier and causes inflammation in the periphery that impacts the central nervous system (CNS) downstream. The meningeal lymphatics are a critical component that drain cerebrospinal fluid from the CNS to the cervical lymph nodes for immunosurveillence. To develop new tools for understanding chemotherapy-mediated effects on the meningeal lymphatics, we present two novel models that examine cellular and tissue level changes. Our in vitro tissue engineered model of a meningeal lymphatic vessel lumen, using a simple tissue culture insert system with both lymphatic endothelial and meningeal cells, examines cell disruption. Our ex vivo model culturing mouse meningeal layers probes structural changes and remodeling, correlating to an explant tissue level. To gain a holistic understanding, we compare our in vitro and ex vivo models to in vivo studies for validation and a three-tier methodology for examining the chemotherapeutic response of the meningeal lymphatics. We have demonstrated that the meningeal lymphatics can be disrupted by systemic chemotherapy but show differential responses to platinum and taxane chemotherapies, emphasizing the need for further study of off-target impacts in the CNS.

Advancing our understanding of skeletal muscle across the lifecourse: Protocol for the MASS_Lifecourse study and characteristics of the first 80 participants.

INTRODUCTION: Sarcopenia, the age-related loss of skeletal muscle strength and mass, carries a significant burden for affected individuals. There has been little investigation of sarcopenia using experimental medicine techniques to study human muscle tissue in detail. The aim of the Muscle Ageing Sarcopenia Studies Lifecourse (MASS_Lifecourse) study is to recruit up to 160 participants, equally divided between females and males between ages 45 and 85 years for detailed phenotyping of skeletal muscle health. Here we describe the protocol for the study and the characteristics of the first 80 participants. METHODS: We are recruiting participants from three sources in the north-east of England. Study fieldwork comprises a home visit (or videocall) for consent and assessment of health, cognition, lifestyle, and wellbeing. This is followed by a visit to a clinical research facility for assessment of sarcopenia status and collection of samples including a vastus lateralis muscle biopsy. We produced descriptive statistics for the first 80 participants, including expressing their grip strength relative to normative data in the form of Z-scores. RESULTS: The first 80 participants (53.8 % female) covered the target ages, ranging from 48 to 84 years. They were regularly physically active, reported good physical function and had a prevalence of sarcopenia (including probable sarcopenia) of 11.3 % based on the revised European consensus. Their grip strength was similar to that in the general population, with a mean Z-score of 0.09 standard deviations (95 % CI: -1.64, 1.83) above that expected. CONCLUSIONS: The MASS_Lifecourse study combines comprehensive health and lifestyle data with a range of biological samples including skeletal muscle. The findings from planned analyses should contribute to improvements in the diagnosis, treatment, and prevention of sarcopenia.

Exit strategies: optimising feasible surveillance for detection, elimination, and ongoing prevention of COVID-19 community transmission.

BACKGROUND: Following implementation of strong containment measures, several countries and regions have low detectable community transmission of COVID-19. We developed an efficient, rapid, and scalable surveillance strategy to detect remaining COVID-19 community cases through exhaustive identification of every active transmission chain. We identified measures to enable early detection and effective management of any reintroduction of transmission once containment measures are lifted to ensure strong containment measures do not require reinstatement. METHODS: We compared efficiency and sensitivity to detect community transmission chains through testing of the following: hospital cases; fever, cough and/or ARI testing at community/primary care; and asymptomatic testing; using surveillance evaluation methods and mathematical modelling, varying testing capacities, reproductive number (R) and weekly cumulative incidence of COVID-19 and non-COVID-19 respiratory symptoms using data from Australia. We assessed system requirements to identify all transmission chains and follow up all cases and primary contacts within each chain, per million population. RESULTS: Assuming 20% of cases are asymptomatic and 30% of symptomatic COVID-19 cases present for testing, with R = 2.2, a median of 14 unrecognised community cases (8 infectious) occur when a transmission chain is identified through hospital surveillance versus 7 unrecognised cases (4 infectious) through community-based surveillance. The 7 unrecognised community upstream cases are estimated to generate a further 55-77 primary contacts requiring follow-up. The unrecognised community cases rise to 10 if 50% of cases are asymptomatic. Screening asymptomatic community members cannot exhaustively identify all cases under any of the scenarios assessed. The most important determinant of testing requirements for symptomatic screening is levels of non-COVID-19 respiratory illness. If 4% of the community have respiratory symptoms, and 1% of those with symptoms have COVID-19, exhaustive symptomatic screening requires approximately 11,600 tests/million population using 1/4 pooling, with 98% of cases detected (2% missed), given 99.9% sensitivity. Even with a drop in sensitivity to 70%, pooling was more effective at detecting cases than individual testing under all scenarios examined. CONCLUSIONS: Screening all acute respiratory disease in the community, in combination with exhaustive and meticulous case and contact identification and management, enables appropriate early detection and elimination of COVID-19 community transmission. An important component is identification, testing, and management of all contacts, including upstream contacts (i.e. potential sources of infection for identified cases, and their related transmission chains). Pooling allows increased case detection when testing capacity is limited, even given reduced test sensitivity. Critical to the effectiveness of all aspects of surveillance is appropriate community engagement, messaging to optimise testing uptake and compliance with other measures.

Mitochondrial Isolevuglandins Contribute to Vascular Oxidative Stress and Mitochondria-Targeted Scavenger of Isolevuglandins Reduces Mitochondrial Dysfunction and Hypertension.

Hypertension remains a major health problem in Western Societies, and blood pressure is poorly controlled in a third of patients despite use of multiple drugs. Mitochondrial dysfunction contributes to hypertension, and mitochondria-targeted agents can potentially improve treatment of hypertension. We have proposed that mitochondrial oxidative stress produces reactive dicarbonyl lipid peroxidation products, isolevuglandins, and that scavenging of mitochondrial isolevuglandins improves vascular function and reduces hypertension. To test this hypothesis, we have studied the accumulation of mitochondrial isolevuglandins-protein adducts in patients with essential hypertension and Ang II (angiotensin II) model of hypertension using mass spectrometry and Western blot analysis. The therapeutic potential of targeting mitochondrial isolevuglandins was tested by the novel mitochondria-targeted isolevuglandin scavenger, mito2HOBA. Mitochondrial isolevuglandins in arterioles from hypertensive patients were 250% greater than in arterioles from normotensive subjects, and ex vivo mito2HOBA treatment of arterioles from hypertensive subjects increased deacetylation of a key mitochondrial antioxidant, SOD2 (superoxide dismutase 2). In human aortic endothelial cells stimulated with Ang II plus TNF (tumor necrosis factor)-α, mito2HOBA reduced mitochondrial superoxide and cardiolipin oxidation, a specific marker of mitochondrial oxidative stress. In Ang II-infused mice, mito2HOBA diminished mitochondrial isolevuglandins-protein adducts, raised Sirt3 (sirtuin 3) mitochondrial deacetylase activity, reduced vascular superoxide, increased endothelial nitric oxide, improved endothelium-dependent relaxation, and attenuated hypertension. Mito2HOBA preserved mitochondrial respiration, protected ATP production, and reduced mitochondrial permeability pore opening in Ang II-infused mice. These data support the role of mitochondrial isolevuglandins in endothelial dysfunction and hypertension. We conclude that scavenging of mitochondrial isolevuglandins may have therapeutic potential in treatment of vascular dysfunction and hypertension.

Scavenging of reactive dicarbonyls with 2-hydroxybenzylamine reduces atherosclerosis in hypercholesterolemic Ldlr-/- mice.

Lipid peroxidation generates reactive dicarbonyls including isolevuglandins (IsoLGs) and malondialdehyde (MDA) that covalently modify proteins. Humans with familial hypercholesterolemia (FH) have increased lipoprotein dicarbonyl adducts and dysfunctional HDL. We investigate the impact of the dicarbonyl scavenger, 2-hydroxybenzylamine (2-HOBA) on HDL function and atherosclerosis in Ldlr-/- mice, a model of FH. Compared to hypercholesterolemic Ldlr-/- mice treated with vehicle or 4-HOBA, a nonreactive analogue, 2-HOBA decreases atherosclerosis by 60% in en face aortas, without changing plasma cholesterol. Ldlr-/- mice treated with 2-HOBA have reduced MDA-LDL and MDA-HDL levels, and their HDL display increased capacity to reduce macrophage cholesterol. Importantly, 2-HOBA reduces the MDA- and IsoLG-lysyl content in atherosclerotic aortas versus 4-HOBA. Furthermore, 2-HOBA reduces inflammation and plaque apoptotic cells and promotes efferocytosis and features of stable plaques. Dicarbonyl scavenging with 2-HOBA has multiple atheroprotective effects in a murine FH model, supporting its potential as a therapeutic approach for atherosclerotic cardiovascular disease.

The role of fit-for-purpose assays within tiered testing approaches: A case study evaluating prioritized estrogen-active compounds in an in vitro human uterotrophic assay.

Chemical risk assessment relies on toxicity tests that require significant numbers of animals, time and costs. For the >30,000 chemicals in commerce, the current scale of animal testing is insufficient to address chemical safety concerns as regulatory and product stewardship considerations evolve to require more comprehensive understanding of potential biological effects, conditions of use, and associated exposures. We demonstrate the use of a multi-level new approach methodology (NAMs) strategy for hazard- and risk-based prioritization to reduce animal testing. A Level 1/2 chemical prioritization based on estrogen receptor (ER) activity and metabolic activation using ToxCast data was used to select 112 chemicals for testing in a Level 3 human uterine cell estrogen response assay (IKA assay). The Level 3 data were coupled with quantitative in vitro to in vivo extrapolation (Q-IVIVE) to support bioactivity determination (as a surrogate for hazard) in a tissue-specific context. Assay AC50s and Q-IVIVE were used to estimate human equivalent doses (HEDs), and HEDs were compared to rodent uterotrophic assay in vivo-derived points of departure (PODs). For substances active both in vitro and in vivo, IKA assay-derived HEDs were lower or equivalent to in vivo PODs for 19/23 compounds (83%). Activity exposure relationships were calculated, and the IKA assay was as or more protective of human health than the rodent uterotrophic assay for all IKA-positive compounds. This study demonstrates the utility of biologically relevant fit-for-purpose assays and supports the use of a multi-level strategy for chemical risk assessment.

Final Results of the Prospective FH02 Mammographic Surveillance Study of Women Aged 35-39 at Increased Familial Risk of Breast Cancer.

BACKGROUND: Many women who are at increased risk of breast cancer due to a mother or sister diagnosed with breast cancer aged under 40 do not currently qualify for surveillance before 40 years of age. There are almost no available data to assess whether mammography screening aged 35-39 years would be effective in this group, in terms of detection of breast cancer at an early stage or cost effective. METHODS: A cohort screening study (FH02) with annual mammography was devised for women aged 35-39 to assess the sensitivity and screening performance and potential survival of women with identified tumours. FINDINGS: 2899 women were recruited from 12/2006-12/2015. These women underwent 12,086 annual screening mammograms and were followed for 13,365.8 years. A total of 55 breast cancers in 54 women occurred during the study period (one bilateral) with 50 cancers (49 women) (15 CIS) adherent to the screening. Eighty percent (28/35) of invasive cancers were ≤ 2 cm and 80% also lymph node negative. Invasive cancers diagnosed in FH02 were significantly smaller than the comparable (POSH-unscreened prospective) study group (45% (131/293) ≤ 2 cm in POSH vs 80% (28/35) in FH02 p < 0.0001), and were less likely to be lymph-node positive (54% (158/290, 3 unknown) in POSH vs 20% (7/35) in FH02: p = 0.0002. Projected and actual survival were also better than POSH. Overall radiation dose was not higher than in an older screened population at mean dose on study per standard sized breast of 1.5 mGy. INTERPRETATION: Mammography screening aged 35-39 years detects breast cancer at an early stage and is likely to be as effective in reducing mortality as in women at increased breast cancer risk aged 40-49 years.

Hepatocellular carcinoma: Epidemiology, pathogenesis and surveillance - implications for sub-Saharan Africa.

Hepatocellular carcinoma (HCC) originates from hepatocytes usually secondary to chronic inflammation and cirrhosis. It is an important disease of global significance with a high incidence and mortality. It is the fifth and eighth most common cancer in males and females, respectively. HCC is also extremely lethal; in 2015 it was the second and sixth most common cause of death from cancer in males and females, respectively. Chronic viral hepatitis B and C are the most frequent risk factors for the development of HCC, and the global distribution of HCC largely mirrors that of chronic viral hepatitis. More recently, there has been a notable increase in the incidence of HCC as a result of obesity-related fatty liver disease. Here, we review the epidemiology of HCC, examine recent advances in our understanding of the pathogenesis of HCC, discuss the implications for identification of potential therapeutic targets, and provide the most updated recommendations on surveillance for HCC, with particular attention to the unique challenges and potential opportunities to reduce the burden of illness and death from HCC in sub-Saharan Africa.

Molecular and cellular oncogenic mechanisms in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC), as the fifth most diagnosed cancer in the world and the third leading cause of death, is a global health concern. Research stimulated by the dismal prognosis of HCC has led to significant advances in the understanding of its aetio-pathogenesis. Dysregulation of genetic, epigenetic and signalling pathways as well as tumour immunological escape mechanisms are implicated in the development of HCC. This review summarises the current knowledge of these mechanisms and argues that it is only through further understanding of their role in hepatocarcinogenesis, that new effective therapies can be developed.

Anti-viral therapy is associated with improved survival but is underutilised in patients with hepatitis B virus-related hepatocellular carcinoma: real-world east and west experience.

BACKGROUND: Hepatitis B virus (HBV) is the leading cause of hepatocellular carcinoma (HCC) worldwide. It remains incompletely understood in the real world how anti-viral therapy affects survival after HCC diagnosis. METHODS: This was an international multicentre cohort study of 2518 HBV-related HCC cases diagnosed between 2000 and 2015. Cox proportional hazards models were utilised to estimate hazard ratios (HR) with 95% (CI) for anti-viral therapy and cirrhosis on patients' risk of death. RESULTS: Approximately, 48% of patients received anti-viral therapy at any time, but only 17% were on therapy at HCC diagnosis (38% at US centres, 11% at Asian centres). Anti-viral therapy would have been indicated for >60% of the patients not on anti-viral therapy based on American criteria. Patients with cirrhosis had lower 5-year survival (34% vs 46%; P < 0.001) while patients receiving anti-viral therapy had increased 5-year survival compared to untreated patients (42% vs 25% with cirrhosis and 58% vs 36% without cirrhosis; P < 0.001 for both). Similar findings were seen for other patient subgroups by cancer stages and cancer treatment types. Anti-viral therapy was associated with a decrease in risk of death, whether started before or after HCC diagnosis (adjusted HR 0.62 and 0.79, respectively; P < 0.001). CONCLUSIONS: Anti-viral therapy improved overall survival in patients with HBV-related HCC across cancer stages and treatment types but was underutilised at both US and Asia centres. Expanded use of anti-viral therapy in HBV-related HCC and better linkage-to-care for HBV patients are needed.

Managing direct oral anticoagulants in patients undergoing dentoalveolar surgery.

Our objective was to describe our experience of managing a cohort of adult patients prescribed direct oral anticoagulants (DOACs) undergoing dentoalveolar procedures between November 2012 and May 2016. Prior to conducting a procedure a formal assessment was made of each patient's anticoagulation treatment. A specific plan was then formulated, balancing the risk of bleeding with the risk of thrombosis. Patients received a telephone consultation one week following treatment to assess any post-operative bleeding. Eighty-two patients underwent 111 oral surgical procedures, the majority of which were dental extractions. In the case of 35 (32%) procedures, advice was given to omit the DOAC, either before or after treatment. There was no bleeding following the majority of procedures. Persistent bleeding followed 15 (13.5%) procedures, of which 7 (6.3%) procedures required specific intervention. The majority of patients prescribed DOACs can undergo dentoalveolar procedures safely. Important considerations when planning treatment are: (i) when the patient usually takes their dose of DOAC, (ii) the time the procedure is performed and, (iii) when the DOAC is taken post-procedure. In our experience, if these factors are considered carefully, omission of DOAC doses is unlikely to be required for most patients.

Isolevuglandins as a gauge of lipid peroxidation in human tumors.

The cellular production of free radicals or reactive oxygen species (ROS) can lead to protein, lipid or DNA modifications and tumor formation. The cellular lipids undergo structural changes through the actions of enzymes (e.g. cyclooxygenases) or free radicals to form a class of compounds called Isolevuglandins (IsoLGs). The recruitment and continued exposure of tissue to ROS and IsoLGs causes increased cell proliferation, mutagenesis, loss of normal cell function and angiogenesis. The elevated concentration of ROS in cancerous tissues suggests that these mediators play an important role in cancer development. We hypothesized that tumors with elevated ROS levels would similarly possess an increased concentration of IsoLGs when compared with normal tissue. Using D11, an ScFv recombinant antibody specific for IsoLGs, we utilized immunohistochemistry to visualize the presence of IsoLG in human tumors compared to normal adjacent tissue (NAT) to the same tumor. We found that IsoLG concentrations were elevated in human breast, colon, kidney, liver, lung, pancreatic and tongue tumor cells when compared to NAT and believe that IsoLGs can be used as a gauge indicative of lipid peroxidation in tumors.

Intraoperative cerebral oxygenation, oxidative injury, and delirium following cardiac surgery.

BACKGROUND: Delirium affects 20-30% of patients after cardiac surgery and is associated with increased mortality and persistent cognitive decline. Hyperoxic reperfusion of ischemic tissues increases oxidative injury, but oxygen administration remains high during cardiac surgery. We tested the hypothesis that intraoperative hyperoxic cerebral reperfusion is associated with increased postoperative delirium and that oxidative injury mediates this association. METHODS: We prospectively measured cerebral oxygenation with bilateral oximetry monitors in 310 cardiac surgery patients, quantified intraoperative hyperoxic cerebral reperfusion by measuring the magnitude of cerebral oxygenation above baseline after any ischemic event, and assessed patients for delirium twice daily in the ICU following surgery using the confusion assessment method for ICU (CAM-ICU). We examined the association between hyperoxic cerebral reperfusion and postoperative delirium, adjusted for the extent of cerebral hypoxia, the extent of cerebral hyperoxia prior to any ischemia, and additional potential confounders and risk factors for delirium. To assess oxidative injury mediation, we examined the association between hyperoxic cerebral reperfusion and delirium after further adjusting for plasma levels of F2-isoprostanes and isofurans at baseline and ICU admission, the association between hyperoxic cerebral reperfusion and these markers of oxidative injury, and the association between these markers and delirium. RESULTS: Ninety of the 310 patients developed delirium following surgery. Every 10%·hour of intraoperative hyperoxic cerebral reperfusion was independently associated with a 65% increase in the odds of delirium (OR, 1.65 [95% CI, 1.12-2.44]; P=0.01). Hyperoxia prior to ischemia was also independently associated with delirium (1.10 [1.01-1.19]; P=0.02), but hypoxia was not (1.12 [0.97-1.29]; P=0.11). Increased hyperoxic cerebral reperfusion was associated with increased concentrations of F2-isoprostanes and isofurans at ICU admission, increased concentrations of these markers were associated with increased delirium, and the association between hyperoxic cerebral reperfusion and delirium was weaker after adjusting for these markers of oxidative injury. CONCLUSIONS: Intraoperative hyperoxic cerebral reperfusion was associated with increased postoperative delirium, and increased oxidative injury following hyperoxic cerebral reperfusion may partially mediate this association. Further research is needed to assess the potential deleterious role of cerebral hyper-oxygenation during surgery.

Novel Deleterious Dihydropyrimidine Dehydrogenase Variants May Contribute to 5-Fluorouracil Sensitivity in an East African Population.

Clinical studies have identified specific genetic variants in dihydropyrimidine dehydrogenase (DPD; DPYD gene) as predictors of severe adverse toxicity to the commonly used chemotherapeutic 5-fluorouracil (5-FU); however, these studies have focused on European and European-American populations. Our laboratory recently demonstrated that additional variants in non-European haplotypes are predictive of 5-FU toxicity. The objective of this study was to identify potential risk variants in an understudied East African population relevant to our institution's catchment area. The DPYD protein-coding region was sequenced in 588 individuals of Somali or Kenyan ancestry living in central/southeast Minnesota. Twelve novel nonsynonymous variants were identified, seven of which significantly decreased DPD activity in vitro. The commonly reported toxicity-associated variants, *2A, D949V, and I560S, were not detected in any individuals. Overall, this study demonstrates a critical limitation in our knowledge of pharmacogenetic predictors of 5-FU toxicity, which has been based on clinical studies conducted in populations of limited diversity.

Scavengers of reactive γ-ketoaldehydes extend Caenorhabditis elegans lifespan and healthspan through protein-level interactions with SIR-2.1 and ETS-7.

Isoketals (IsoKs) are highly reactive γ-ketoaldehyde products of lipid peroxidation that covalently adduct lysine side chains in proteins, impairing their function. Using C. elegans as a model organism, we sought to test the hypothesis that IsoKs contribute to molecular aging through adduction and inactivation of specific protein targets, and that this process can be abrogated using salicylamine (SA), a selective IsoK scavenger. Treatment with SA extends adult nematode longevity by nearly 56% and prevents multiple deleterious age-related biochemical and functional changes. Testing of a variety of molecular targets for SA's action revealed the sirtuin SIR-2.1 as the leading candidate. When SA was administered to a SIR-2.1 knockout strain, the effects on lifespan and healthspan extension were abolished. The SIR-2.1-dependent effects of SA were not mediated by large changes in gene expression programs or by significant changes in mitochondrial function. However, expression array analysis did show SA-dependent regulation of the transcription factor ets-7 and associated genes. In ets-7 knockout worms, SA's longevity effects were abolished, similar to sir-2.1 knockouts. However, SA dose-dependently increases ets-7 mRNA levels in non-functional SIR-2.1 mutant, suggesting that both are necessary for SA's complete lifespan and healthspan extension.

Controlled moderate hypovolaemia in healthy volunteers is not associated with the development of oxidative stress assessed by plasma F2-isoprostanes and isofurans.

Hypovolaemia can be associated with substantial morbidity, particularly when it occurs in the setting of trauma and in patients with comorbid diseases. Hypovolaemia and inflammation such as occur in the setting of trauma and surgery, are associated with systemic oxidative stress and free-radical injury. Free-radical injury that results from hypovolaemia-induced organ reperfusion may further augment inflammatory processes. It is unknown exactly what proportion of free-radical injury is associated with isolated hypovolaemia as opposed to the contribution from inflammation from surgery or trauma. In the first human study of its kind, we exposed 8 adult male volunteers to venesection-induced hypovolaemia in progressive aliquots of 5% of total blood volume until 20% had been removed. This blood was subsequently reinfused. Plasma F2-isoprostanes and isofurans, markers of in vivo lipid oxidation, were measured by gas chromatography-mass spectrometry at each 5% aliquot venesected and at each 5% reinfused. Between baseline and maximal blood loss there was a minor fall in haemoglobin concentration from 143.9g/l to 138.8g/l (p=0.004, 95% CI 2.2, 8.0g/L). No significant change from baseline occurred in the concentrations of either plasma F2-isoprostanes or isofurans during venesection (p=0.116 and p=0.152, respectively) or blood reinfusion (p=0.553 and p=0.736, respectively). We can conclude that in healthy adult volunteers, isolated hypovolaemia to 20% total blood volume loss is not associated with detectable systemic oxidative stress. The free-radical injury identified in surgical and trauma patients may represent the effects of tissue damage and inflammation, with an uncertain contribution from tissue ischemia as may occur with hypovolaemia.

Mitochondrial dysfunction and oxidative stress in patients with chronic kidney disease.

Mitochondria abnormalities in skeletal muscle may contribute to frailty and sarcopenia, commonly present in patients with chronic kidney disease (CKD). Dysfunctional mitochondria are also a major source of oxidative stress and may contribute to cardiovascular disease in CKD We tested the hypothesis that mitochondrial structure and function worsens with the severity of CKD Mitochondrial volume density, mitochondrial DNA (mtDNA) copy number, BNIP3, and PGC1α protein expression were evaluated in skeletal muscle biopsies obtained from 27 subjects (17 controls and 10 with CKD stage 5 on hemodialysis). We also measured mtDNA copy number in peripheral blood mononuclear cells (PBMCs), plasma isofurans, and plasma F2-isoprostanes in 208 subjects divided into three groups: non-CKD (eGFR>60 mL/min), CKD stage 3-4 (eGFR 60-15 mL/min), and CKD stage 5 (on hemodialysis). Muscle biopsies from patients with CKD stage 5 revealed lower mitochondrial volume density, lower mtDNA copy number, and higher BNIP3 content than controls. mtDNA copy number in PBMCs was decreased with increasing severity of CKD: non-CKD (6.48, 95% CI 4.49-8.46), CKD stage 3-4 (3.30, 95% CI 0.85-5.75, P = 0.048 vs. non-CKD), and CKD stage 5 (1.93, 95% CI 0.27-3.59, P = 0.001 vs. non-CKD). Isofurans were higher in patients with CKD stage 5 (median 59.21 pg/mL, IQR 41.76-95.36) compared to patients with non-CKD (median 49.95 pg/mL, IQR 27.88-83.46, P = 0.001), whereas F2-isoprostanes did not differ among groups. Severity of CKD is associated with mitochondrial dysfunction and markers of oxidative stress. Mitochondrial abnormalities, which are common in skeletal muscle from patients with CKD stage 5, may explain the muscle dysfunction associated with frailty and sarcopenia in CKD Further studies are required to evaluate mitochondrial function in vivo in patients with different CKD stages.

The relationship between plasma lipid peroxidation products and primary graft dysfunction after lung transplantation is modified by donor smoking and reperfusion hyperoxia.

BACKGROUND: Donor smoking history and higher fraction of inspired oxygen (FIO2) at reperfusion are associated with primary graft dysfunction (PGD) after lung transplantation. We hypothesized that oxidative injury biomarkers would be elevated in PGD, with higher levels associated with donor exposure to cigarette smoke and recipient hyperoxia at reperfusion. METHODS: We performed a nested case-control study of 72 lung transplant recipients from the Lung Transplant Outcomes Group cohort. Using mass spectroscopy, F2-isoprostanes and isofurans were measured in plasma collected after transplantation. Cases were defined in 2 ways: grade 3 PGD present at day 2 or day 3 after reperfusion (severe PGD) or any grade 3 PGD (any PGD). RESULTS: There were 31 severe PGD cases with 41 controls and 35 any PGD cases with 37 controls. Plasma F2-isoprostane levels were higher in severe PGD cases compared with controls (28.6 pg/ml vs 19.8 pg/ml, p = 0.03). Plasma F2-isoprostane levels were higher in severe PGD cases compared with controls (29.6 pg/ml vs 19.0 pg/ml, p = 0.03) among patients reperfused with FIO2 >40%. Among recipients of lungs from donors with smoke exposure, plasma F2-isoprostane (38.2 pg/ml vs 22.5 pg/ml, p = 0.046) and isofuran (66.9 pg/ml vs 34.6 pg/ml, p = 0.046) levels were higher in severe PGD compared with control subjects. CONCLUSIONS: Plasma levels of lipid peroxidation products are higher in patients with severe PGD, in recipients of lungs from donors with smoke exposure, and in recipients exposed to higher Fio2 at reperfusion. Oxidative injury is an important mechanism of PGD and may be magnified by donor exposure to cigarette smoke and hyperoxia at reperfusion.