Proximal humeral osteoarticular allografts: technique,pearls and pitfalls,outcomes

Allograft transplantation is a biologic reconstruction option for massive bone defects after resection of bone sarcomas. This type of reconstruction not only restores bone stock but it also allows us to reconstruct the joint anatomically. These factors are a major concern, especially in a young and active population.We are describing indications, surgical techniques, pearls and pitfalls, and outcomes of proximal humeral osteoarticular allografts, done at present time in our institution.We found that allograft fractures and articular complications, as epiphyseal resorption and subchondral fracture, are the main complications observed in proximal humerus osteoarticular allograft reconstructions. Nevertheless, only fractures need a reconstruction revision. Joint complications may adversely affect the limb function, but for this reason, an allograft revision is rarely performed.     

ATG16L1 and NOD2 polymorphisms enhance phagocytosis in monocytes of Crohn’s disease patients

AIM:To investigate if the presence of relevant genetic polymorphisms has effect on the effectual clearance of bacteria by monocytes and granulocytes in patients with Crohn’s disease(CD).METHODS:In this study,we assessed the differential responses in phagocytosis by measuring the phagocytic activity and the percentage of active phagocytic monocytes and granulocytes in inflammatory bowel disease patients as well as healthy controls.As both autophagy related like 1(ATG16L1)and immunityrelated guanosine triphosphatase gene are autophagy genes associated with CD and more recently nucleo-tide-binding ligomerization domain-containing protein2(NOD2)has been identified as a potent inducer of autophagy we genotyped the patients for these variants and correlated this to the phagocytic reaction.The genotyping was done with restriction fragment length polymorphisms analysis and the phagocytosis was determined with the pHrodo?Escherichia coli Bioparticles Phagocytosis kit for flowcytometry.RESULTS:In this study,we demonstrate that analysis of the monocyte and granulocyte populations of patients with CD and ulcerative colitis showed a comparable phagocytic activity(ratio of mean fluorescence intensity)between the patient groups and the healthy controls.CD patients show a significantly higher phagocytic capacity(ratio mean percentage of phagocytic cells)compared to healthy controls(51.91%±2.85%vs 37.67%±7.06%,P=0.05).The extend of disease was not of influence.However,variants of ATG16L1(WT:2.03±0.19 vs homozygoot variant:4.38±0.37,P<0.009)as well as NOD2(C-ins)(heterozygous variant:42.08±2.94 vs homozygous variant:75.58±4.34(P=0.05)are associated with the phagocytic activity in patients with CD.CONCLUSION:Monocytes of CD patients show enhanced phagocytosis associated with the presence of ATG16L1 and NOD2 variants.This could be part of the pathophysiological mechanism resulting in the disease.     

Metformin and phenformin deplete tricarboxylic acid cycle and glycolytic intermediates during cell transformation and NTPs in cancer stem cells

Metformin, a first-line diabetes drug linked to cancer prevention in retrospective clinical analyses, inhibits cellular transformation and selectively kills breast cancer stem cells (CSCs). Although a few metabolic effects of metformin and the related biguanide phenformin have been investigated in established cancer cell lines, the global metabolic impact of biguanides during the process of neoplastic transformation and in CSCs is unknown. Here, we use LC/MS/MS metabolomics (>200 metabolites) to assess metabolic changes induced by metformin and phenformin in an Src-inducible model of cellular transformation and in mammosphere-derived breast CSCs. Although phenformin is the more potent biguanide in both systems, the metabolic profiles of these drugs are remarkably similar, although not identical. During the process of cellular transformation, biguanide treatment prevents the boost in glycolytic intermediates at a specific stage of the pathway and coordinately decreases tricarboxylic acid (TCA) cycle intermediates. In contrast, in breast CSCs, biguanides have a modest effect on glycolytic and TCA cycle intermediates, but they strongly deplete nucleotide triphosphates and may impede nucleotide synthesis. These metabolic profiles are consistent with the idea that biguanides inhibit mitochondrial complex 1, but they indicate that their metabolic effects differ depending on the stage of cellular transformation.Altered metabolism is a hallmark of malignantly transformed cells. Cancer risk is linked to metabolic syndrome, a disease state that includes obesity, type 2 diabetes, high cholesterol, and atherosclerosis. Retrospective studies of type 2 diabetes patients treated with metformin, the most widely prescribed antidiabetic drug, show a strong correlation between drug intake and reduced tumor incidence or reduced cancer-related deaths (1–4).In the breast lineage, metformin inhibits growth of cancer cell lines (5–7), blocks transformation in a Src-inducible cell system (8, 9), and selectively inhibits the growth of cancer stem cells (CSCs) (8). As a consequence of its selective effects on CSCs, combinatorial therapy of metformin and standard chemotherapeutic drugs (doxorubicin, paclitaxel, and cisplatin) increases tumor regression and prolongs remission in mouse xenografts (8, 10). In addition, metformin can decrease the chemotherapeutic dose for prolonging tumor remission in xenografts involving multiple cancer types (10).Phenformin, a related biguanide and formerly used diabetes drug, acts as an anticancer agent in tumors including lung, lymphoma, and breast cancer with a greater potency than metformin. Phenformin mediates antineoplastic effects at a lower concentration than metformin in cell lines, a PTEN-deficient mouse model, breast cancer xenografts, and drug-induced mitochondrial impairment (11–14). The chemical similarities of these biguanides, as well as their similar effects in diabetes and cancer, have led to the untested assumption that phenformin is essentially a stronger version of metformin.In a Src-inducible model of cellular transformation and CSC formation, multiple lines of evidence suggest that metformin inhibits a signal transduction pathway that results in an inflammatory response (15). In the context of atherosclerosis, metformin inhibits NF-κB activation and the inflammatory response via a pathway involving AMP kinase (AMPK) and the tumor suppressor PTEN (16, 17). As metformin alters energy metabolism in diabetics, we speculated that metformin might block a metabolic stress response that stimulates the inflammatory pathway (15). However, very little is known about the metabolic changes that inhibit the inflammatory pathway.Previous studies on metformin-induced metabolic effects in cancer have focused on single metabolic alterations or pathways in already established cancer cell lines. Metformin leads to activation of AMPK, which plays a key role in insulin signaling and energy sensing (18). Metformin can reduce protein synthesis via mTOR inhibition (19). In addition, metformin may directly impair mitochondrial respiration through complex I inhibition and has been described to boost glycolysis as a compensation mechanism (14, 20). In this regard, lactic acidosis can be a side effect of metformin and phenformin treatment of diabetic patients, presumably because inhibition of complex I prevents NADH oxidation, thereby leading to a requirement for cytosolic NADH to be oxidized by the conversion of pyruvate to lactate. There is some knowledge about the metabolic effects of metformin (21, 22), but very little is known about the specific metabolic alterations linking biguanides to inhibition of neoplastic transformation.Here, we perform a metabolomic analysis on the effects of metformin and phenformin in a Src-inducible model of transformation and in CSCs. This inducible model permits an analysis of the transition from nontransformed to transformed cells in an isogenic cell system and hence differs from analyses of already established cancer cell lines. We studied CSCs to address why this population, which is resistant to standard chemotherapeutics and hypothesized to be a major reason for tumor recurrence, is selectively inhibited by metformin. Our results indicate the metabolic effects of metformin and phenformin are remarkably similar to each other, with only a few differences. Both biguanides dramatically decrease tricarboxylic acid (TCA) cycle intermediates in the early stages of transformation, and they inhibit the boost in select glycolytic intermediates that normally occurs with transformation along with increases in glycerol 3-phosphate and lactate, which are metabolites branching from glycolysis. Unexpectedly, in CSCs, biguanides have only marginal effects on glycolytic and TCA cycle metabolites, but they severely decrease nucleotide triphosphates. These detailed metabolic analyses provide independent support for the idea that metformin inhibits mitochondrial complex 1 (14, 20), and they indicate that the metabolic effects of biguanides depend on the stage of the cellular transformation.     

A common X-linked inborn error of carnitine biosynthesis may be a risk factor for nondysmorphic autism

We recently reported a deletion of exon 2 of the trimethyllysine hydroxylase epsilon (TMLHE) gene in a proband with autism. TMLHE maps to the X chromosome and encodes the first enzyme in carnitine biosynthesis, 6-N-trimethyllysine dioxygenase. Deletion of exon 2 of TMLHE causes enzyme deficiency, resulting in increased substrate concentration (6-N-trimethyllysine) and decreased product levels (3-hydroxy-6-N-trimethyllysine and γ-butyrobetaine) in plasma and urine. TMLHE deficiency is common in control males (24 in 8,787 or 1 in 366) and was not significantly increased in frequency in probands from simplex autism families (9 in 2,904 or 1 in 323). However, it was 2.82-fold more frequent in probands from male-male multiplex autism families compared with controls (7 in 909 or 1 in 130; P = 0.023). Additionally, six of seven autistic male siblings of probands in male-male multiplex families had the deletion, suggesting that TMLHE deficiency is a risk factor for autism (metaanalysis Z-score = 2.90 and P = 0.0037), although with low penetrance (2-4%). These data suggest that dysregulation of carnitine metabolism may be important in nondysmorphic autism; that abnormalities of carnitine intake, loss, transport, or synthesis may be important in a larger fraction of nondysmorphic autism cases; and that the carnitine pathway may provide a novel target for therapy or prevention of autism.