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Signaling from multiple receptor tyrosine kinases (RTK) contributes to therapeutic resistance in glioblastoma (GBM). Heparan sulfate (HS), present on cell surfaces and in the extracellular matrix, regulates cell signaling via several mechanisms. To investigate the role for HS in promoting RTK signaling in GBM, we generated neural progenitor cells deficient for HS by knockout of the essential HS-biosynthetic enzyme Ext1, and studied tumor initiation and progression. HS-null cells had decreased proliferation, invasion, and reduced activation of multiple RTKs compared with control. In vivo tumor establishment was significantly decreased, and rate of tumor growth reduced with HS-deficient cells implanted in an HS-poor microenvironment. To investigate if HS regulates RTK activation through platelet-derived growth factor receptor α (PDGFRα) signaling, we removed cell surface HS in patient-derived GBM lines and identified reduced cell surface PDGF-BB ligand. Reduced ligand levels were associated with decreased phosphorylation of PDGFRα, suggesting HS promotes ligand–receptor interaction. Using human GBM tumorspheres and a murine GBM model, we show that ligand-mediated signaling can partially rescue cells from targeted RTK inhibition and that this effect is regulated by HS. Indeed, tumor cells deficient for HS had increased sensitivity to EGFR inhibition in vitro and in vivo.Implications:Our study shows that HS expressed on tumor cells and in the tumor microenvironment regulates ligand-mediated signaling, promoting tumor cell proliferation and invasion, and these factors contribute to decreased tumor cell response to targeted RTK inhibition.

Glypican-1 and Syndecan-1 are identified as heparan sulfate proteoglycans expressed on GBM43.

Cotoneaster integerrimus represents a multiploid and facultative apomictic system of widely distributed mountain populations. We used flow cytometry to determine genome size, ploidy level, and reproduction mode variation of the Balkan populations, supplemented by analysis of nuclear microsatellites in order to address: (i) geographic distribution and variation of cytotypes among the populations; (ii) variation of reproduction mode and the frequency of sexuality; (iii) pathways of endosperm formation among the sampled polyploids and their endosperm balance requirements; (iv) genotypic diversity and geographic distribution of clonal lineages of polyploids. The prevalence of apomictic tetraploid cytotype followed by sexual diploids and extremely rare triploids was demonstrated. This prevalence of tetraploids affected the populations’ structure composed from clonal genotypes with varying proportions. The co-occurrence of diploids and tetraploids generated higher cytotype, reproductive mode, and genotypic diversity, but mixed-ploidy sites were extremely rare. The endosperm imbalance facilitates the development and the occurrence of intermediate triploids in mixed-ploidy populations, but also different tetraploid lineages elsewhere with unbalanced endosperm. All these results showed that the South European populations of C. integerrimus have higher levels of cytotype and reproductive diversity compared to the Central European ones. Therefore, the South European populations can be considered as a potential reservoir of regional and global diversity for this species.

Cerebellar Purkinje neurons help compute absolute subsecond timing, but how their firing is affected during repetitive sensory stimulation with consistent subsecond intervals remains unaddressed. Here, we investigated how simple and complex spikes of Purkinje cells change during regular application of air puffs (3.3 Hz for ∼4 min) to the whisker pad of awake, head-fixed female mice. Complex spike responses fell into two categories: those in which firing rates increased (at ∼50 ms) and then fell [complex spike elevated (CxSE) cells] and those in which firing rates decreased (at ∼70 ms) and then rose [complex spike reduced (CxSR) cells]. Both groups had indistinguishable rates of basal complex (∼1.7 Hz) and simple (∼75 Hz) spikes and initially responded to puffs with a well-timed sensory response, consisting of a short-latency (∼15 ms), transient (4 ms) suppression of simple spikes. CxSE more than CxSR cells, however, also showed a longer-latency increase in simple spike rate, previously shown to reflect motor command signals. With repeated puffs, basal simple spike rates dropped greatly in CxSR but not CxSE cells; complex spike rates remained constant, but their temporal precision rose in CxSR cells and fell in CxSE cells. Also over time, transient simple spike suppression gradually disappeared in CxSE cells, suggesting habituation, but remained stable in CxSR cells, suggesting reliable transmission of sensory stimuli. During stimulus omissions, both categories of cells showed complex spike suppression with different latencies. The data indicate two modes by which Purkinje cells transmit regular repetitive stimuli, distinguishable by their climbing fiber signals. NEW & NOTEWORTHY Responses of cerebellar Purkinje cells in awake mice form two categories defined by complex spiking during regular trains of brief, somatosensory stimuli. Cells in which complex spike probability first increases or decreases show simple spike suppressions that habituate or persist, respectively. Stimulus omissions alter complex spiking. The results provide evidence for differential suppression of olivary cells during sensory stimulation and omissions and illustrate that climbing fiber innervation defines Purkinje cell responses to repetitive stimuli.

Voltage-gated Na channels of Purkinje cells are specialized to maintain high availability during high-frequency repetitive firing. They enter fast-inactivated states relatively slowly and undergo a voltage-dependent open-channel block by an intracellular protein (or proteins) that prevents stable fast inactivation and generates resurgent Na current. These properties depend on the pore-forming α subunits, as well as modulatory subunits within the Na channel complex. The identity of the factors responsible for open-channel block remains a question. Here we investigate the effects of genetic mutation of two Na channel auxiliary subunits highly expressed in Purkinje cells, NaVβ4 and FGF14, on modulating Na channel blocked as well as inactivated states. We find that although both NaVβ4 and the FGF14 splice variant FGF14-1a contain sequences that can generate resurgent-like currents when applied to Na channels in peptide form, deletion of either protein, or both proteins simultaneously, does not eliminate resurgent current in acutely dissociated Purkinje cell bodies. Loss of FGF14 expression does, however, reduce resurgent current amplitude and leads to an acceleration and stabilization of inactivation that is not reversed by application of the site-3 toxin, anemone toxin II (ATX). Tetrodotoxin (TTX) sensitivity is higher for resurgent than transient components of Na current, and loss of FGF14 preferentially affects a highly TTX-sensitive subset of Purkinje α subunits. The data suggest that NaV1.6 channels, which are known to generate the majority of Purkinje cell resurgent current, bind TTX with high affinity and are modulated by FGF14 to facilitate open-channel block.

Introduction: Abdominal aortic aneurysms (AAA) are responsible for over 150,000 deaths worldwide annually. Attempts at producing a reliable large-animal model of AAA have proven challenging. We sought to create a reproducible swine model of AAA using enzymatic degradation of the aortic wall. Methods: A total of 9 male Yorkshire swine received periadventitial injections of type 1 collagenase (23.5 mg) and porcine pancreatic elastase (500 mg) into a 4 cm segment of infrarenal aorta. Aortic diameter growth was monitored at POD 7 and 14 using ultrasound. The animals were euthanized on POD 21, and the suprarenal (control) and infrarenal (treated) aorta was harvested for analysis, after gross measurement of aortic diameter under physiologic blood pressure. Sections of control and treated aorta were used to obtain tensile strength using a tensiometer. Additional segments of the aorta were collected for histopathological analysis (H&E, elastin, alpha smooth muscle actin). PCR of matrix metalloproteinases (MMP9) was conducted. Groups were compared with paired t-tests, or ANOVA for repeated measures, where appropriate. Results: Average percent growth of aortic diameter at POD 21 for treated segments was 27% +/- 16.5% versus 4.5% +/- 4% for control tissue. The average difference in aortic growth by subject, was 26.7% [14.6%-38.8%]; (p Conclusion: A reproducible, large-animal model of AAA, with anatomical, histopathological, and biomechanical properties that are clinically translatable, can be achieved with extraluminal enzymatic degradation.

Rhabdomyosarcoma (RMS) and rhabdoid tumors (RT) are rare soft-tissue malignancies with the highest incidence in infants, children, and adolescents. Advanced, recurrent, and/or metastatic RMS and RT exhibit poor response to treatment. One of the main mechanisms behind resistance to treatment is believed to be intratumoral heterogeneity. In this study, we investigated the myogenic determination factor 1 (MYOD1) and Noggin (NOG) markers in an embryonal RMS (ERMS) cell line and an RT cell line and the differential response of the MYOD1 and NOG expressing subpopulations to chemotherapy. Importantly, we found that these markers together identify a subpopulation of cells (MYOD1+ NOG+ cells) with primary resistance to Vincristine and Doxorubicin, two commonly used chemotherapies for ERMS and RT. The chemoresistant MYOD1+ NOG+ cells express markers of undifferentiated cells such as myogenin and ID1. Combination of Vincristine with TPA/GSK126, a drug combination shown to induce differentiation of RMS cell lines, is able to partially overcome MYOD1/NOG cells chemoresistance.

Objective: (1) Determine if sequential administration of standard chemotherapy (paclitaxel and cisplatin, P/C) with epigenetic drugs effectively targets ovarian cancer and limits toxicity to normal cells. (2) Define whether epigenetic treatment can shorten the exposure to P/C. Methods: Normal cells—adipocyte-derived stem cells (ASC), primary fibroblasts (PF), and human intestinal epithelial cells (HIEC-6)—were treated with 48 h IC50 values of P/C and epigenetic drugs, 5-azacytidine (AZA) and or suberoylanilide hydroxamic acid (SAHA), in combination and sequentially. The least toxic regimens to normal cells were administered to the ovarian cancer cell lines Caov-3, SKOV-3, OVCAR-3. Cell viability after treatments were assessed using the 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide (MTT) and cell count assays. Secretome analysis of conditioned medium collected from the treated ovarian cancer cells was performed using ELISA. Results: P/C with AZA and SAHA targeted all ovarian cancer cell lines (82-99% cell death), but also caused significant normal cell death (66-100%). In contrast, P/C followed by AZA or SAHA is less toxic to ASC and PF (25-96% viability) when compared to a four-drug combination therapy (1% viability, p

Flow cytometry has become the method of choice to measure the DNA content (genome size) in plants. Ease of sample preparation, fast acquisition, and accurate measurements have made the method popular in the domains of plant cell biology, systematics, evolution, genetics and biotechnology. Although the cell wall is a problem when isolating plant cells, cytometry remains a powerful tool in plant sciences. Based on our 30-years’ experience in this field, this review will focus at first on genome size measurement using simply isolated nuclei: the good practice for acquisition, nuclei isolation, appropriate buffers, kind of tissues to use. The second part will briefly review what kind of measurements it is possible to make in plant cytometry, and for what purpose: base composition, ploidy level, cell cycle, endoreplication, seed screening, and nuclei/chromosomes sorting. We will address troubleshooting. The commonly-used mathematic tools will be discussed.

Objective Adipose-derived stem cells (ASCs) are a potential adult mesenchymal stem cell source for restoring endothelial function in patients with critical limb ischemia. Fibroblast growth factor 2 (FGF2) and vascular endothelial growth factor (VEGF) play a major role in angiogenesis and wound healing. This study evaluated the effects of FGF and VEGF on the proliferation, migration, and potential endothelial differentiation of human ASCs with regards to their use as endothelial cell substitutes. Methods ASCs were isolated from clinical lipoaspirates and cultured in M199 medium with fetal bovine serum (10%), FGF2 (10 ng/mL), VEGF (50 ng/mL), or combinations of FGF2 and VEGF. Cell proliferation rates, viability, and migration were measured by growth curves, MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide), and scratch assays. For cell attachment determinations, ASCs were seeded onto a scaffold of small intestinal submucosa for 5 days. Endothelial differentiation capabilities of ASCs were confirmed by expression of endothelial cell-specific markers using quantitative polymerase chain reaction, immunofluorescence staining, and cord formation on Matrigel (BD Biosciences, San Jose, Calif). PD173074, a selective inhibitor of FGF receptor, was used to confirm the importance of FGF signaling. Results ASCs treated with FGF or combinations of FGF and VEGF showed increased proliferation rates and consistent differentiation toward an endothelial cell lineage increase in platelet endothelial cell adhesion molecule (CD31), von Willebrand factor, endothelial nitric oxide synthase, and vascular endothelial cadherin message, and in protein and cord formation on Matrigel. FGF and VEGF stimulated ASC migration and increased the attachment and retention after seeding onto a matrix graft of small intestinal submucosa. Blockade of FGF signaling with PD173074 abrogated ASC endothelial cell differentiation potential. Conclusions These results indicate that FGF and VEGF are ASC promoters for proliferation, migration, attachment, and endothelial differentiation. FGF and VEGF have a costimulatory effect on ASC endotheliogenesis. These results further suggest that ASCs with enhanced FGF signaling may potentially be used for tissue engineering and cell-based therapies in patients with critical limb ischemia.

Background aims Adipose-derived stem cells (ASCs) are considered to play a positive role in wound healing as evidenced by their increasing use in breast reconstructive procedures. After chemotherapy for breast cancer, poor soft tissue wound healing is a major problem. In the present study, the functional capabilities and recovery of ASCs after exposure to chemotherapeutic agent paclitaxel (PTX) using in vitro and ex vivo models were demonstrated. Methods Human ASCs were isolated from periumbilical fat tissue and treated with PTX at various concentrations. Adult Sprague-Dawley rats were given intravenous injections with PTX. Two and four weeks after the initial PTX treatment, ASCs were isolated from rat adipose tissue. Proliferation, cell viability, apoptosis and cell migration rates were measured by growth curves, MTT assays, flow cytometry and scratch assays. ASCs were cultured in derivative-specific differentiation media with or without PTX for 3 weeks. Adipogenic, osteogenic and endothelial differentiation levels were measured by quantitative reverse transcriptase polymerase chain reaction and histological staining. Results PTX induced apoptosis, decreased the proliferation and cell migration rates of ASCs and inhibited ASCs multipotent differentiation in both in vitro human ASC populations and ex vivo rat ASC populations with PTX treatment. Furthermore, after cessation of PTX, ASCs exhibited recovery potential of differentiation capacity in both in vitro and animal studies. Conclusions Our results provide insight into poor soft tissue wound healing and promote further understanding of the potential capability of ASCs to serve as a cell source for fat grafting and reconstruction in cancer patients undergoing chemotherapy treatment.

Purkinje neurons express voltage-gated Na channels and auxiliary proteins that facilitate rapid firing. White et al. find that Na current in isolated Purkinje cells is unaffected by loss of NaVβ4 subunits, but transient current is briefer and resurgent current is reduced without the FGF14 protein., Voltage-gated Na channels of Purkinje cells are specialized to maintain high availability during high-frequency repetitive firing. They enter fast-inactivated states relatively slowly and undergo a voltage-dependent open-channel block by an intracellular protein (or proteins) that prevents stable fast inactivation and generates resurgent Na current. These properties depend on the pore-forming α subunits, as well as modulatory subunits within the Na channel complex. The identity of the factors responsible for open-channel block remains a question. Here we investigate the effects of genetic mutation of two Na channel auxiliary subunits highly expressed in Purkinje cells, NaVβ4 and FGF14, on modulating Na channel blocked as well as inactivated states. We find that although both NaVβ4 and the FGF14 splice variant FGF14-1a contain sequences that can generate resurgent-like currents when applied to Na channels in peptide form, deletion of either protein, or both proteins simultaneously, does not eliminate resurgent current in acutely dissociated Purkinje cell bodies. Loss of FGF14 expression does, however, reduce resurgent current amplitude and leads to an acceleration and stabilization of inactivation that is not reversed by application of the site-3 toxin, anemone toxin II (ATX). Tetrodotoxin (TTX) sensitivity is higher for resurgent than transient components of Na current, and loss of FGF14 preferentially affects a highly TTX-sensitive subset of Purkinje α subunits. The data suggest that NaV1.6 channels, which are known to generate the majority of Purkinje cell resurgent current, bind TTX with high affinity and are modulated by FGF14 to facilitate open-channel block.

The existence of two chromosome numbers (2n = 16 and 2n = 40) in Lygeum spartum is confirmed in the Algerian steppe populations of Oran region. Chromosome counts were established for 11 Algerian populations and three supplementary populations from Italy, Spain and Greece. The karyotypes were characterized by chromosome markers obtained using fluorescence in situ hybridization and fluorochrome bandings. The organization of 5S and 18S-5.8S-26S (35S) rDNA was studied in both cytotypes. Six signals of 35S and 2 signals of 5S were observed in 2n = 16 population, while 10 signals of 35S and 4 signals of 5S were detected in the population with 2n = 40. All 35S loci were also strongly marked by chromomycin, but negatively stained by Hoechst, which indicates the presence of GC rich DNA in rDNA regions. The B chromosomes were found in both cytotypes, bearing a 35S locus in 2n = 16 population. Genome size, determined by cytometry of 10 populations, ranged from 9.27 pg for 2n = 16 to 26.63 pg for 2n = 40 populations. The sequencing of plastid and nuclear DNA markers did not reveal major differences among 2n = 16 and 2n = 40 populations. However, given the differences between two cytotypes and based on their morphological and cytogenetic characteristics, the 2n = 16 cytotype merits novel taxonomic treatment., This study was supported by the CMEP (Le Comité Mixte d'Evaluation et de Prospective Franco-Algérien - Programme TASSILI; Nb. 01 MDU/530).

Introduction: We investigated interrater reliability of overall assessment of nerve root lesions by electrodiagnostic testing (EDX) in neonatal brachial plexus palsy (NBPP). Methods: Two blinded, board-certified reviewers retrospectively reviewed de-identified EDX data from 37 infants with NBPP for 2005–2012. Only nerve conduction and electromyography needle data were included. The examiners independently assigned 1 of 4 nerve root lesion categories: (1) pre-ganglionic lesion (avulsion), (2) post-ganglionic lesion (rupture), (3) normal, or (4) “unable to determine.” Simple percentage agreement, the Cohen kappa statistic representing interrater reliability for each nerve root (C5–T1), and overall kappa between examiners were evaluated. Results: Interrater reliabilities were substantial to almost perfect for each nerve root except C5. Considering all nerve roots, overall interrater reliability was substantial (kappa = 0.62); simple percentage agreement was 75% (138/185). Conclusions: Interrater reliability of nerve root assessment by EDX for infants with NBPP was high for C6–T1 root levels, but less reliable for C5 because of technical factors. Muscle Nerve 55: 69–73, 2017

To test how cerebellar crus I/II Purkinje cells and their targets in the lateral cerebellar nuclei (CbN) integrate sensory and motor-related inputs and contribute to reflexive movements, we recorded extracellularly in awake, head-fixed mice during non-contact whisking. Ipsilateral or contralateral air puffs elicited changes in population Purkinje simple spike rates that matched whisking kinematics (∼1 Hz/1° protraction). Responses remained relatively unaffected when ipsilateral sensory feedback was removed by lidocaine but were reduced by optogenetically inhibiting the reticular nuclei. Optogenetically silencing cerebellar output suppressed movements. During puff-evoked whisks, both Purkinje and CbN cells generated well-timed spikes in sequential 2- to 4-ms windows at response onset, such that they alternately elevated their firing rates just before protraction. With spontaneous whisks, which were smaller than puff-evoked whisks, well-timed spikes were absent and CbN cells were inhibited. Thus, sensory input can facilitate millisecond-scale, well-timed spiking in Purkinje and CbN cells and amplify reflexive whisker movements.

Background Adipose-derived stem cells (ASCs) assisted lipotransfer have been considered to facilitate the survival of fat grafts. However, emerging evidence of insufficient vascularization is another obstacle for fat graft survival in cell-assisted lipotransfer. Objectives This study evaluated if endothelial phenotype ASCs with fat lipoaspirate improves survival and neovascularization in fat transplantation Methods ASCs were isolated from human periumbilical fat tissue and cultured in endothelial growth medium for 2 weeks. Fat lipoaspirate was mixed with fresh adipose stroma vascular fraction (SVF), endothelial differentiated ASCs (EC/ASCs), and fat lipoaspirate alone. Three fat mixtures were subcutaneously injected into the adult male Sprague–Dawley rat’s dorsum at 3 locations. At 8 weeks after transplantation, the grafted fat lipoaspirates were harvested, and the extracted fat was evaluated using photographic, survival weights measurements and histological examination. Neo-vascularization was quantified by immunofluorescence and real-time RT-PCR. Results Grafts from the EC/ASC assisted group had a higher survival rate, morphologic integrity, and most uniform lipid droplets. They also revealed less inflammation and fibrosis with increased number of vessels by histological and immunofluorescence analysis. Quantitative RT-PCR analysis indicated that the expression levels of EC-specific markers of CD31 and vWF were higher in the EC/ASC group compared with in the control and fat with SVF transplants. Conclusions These results indicated that co-implantation of fat lipoaspirate with ASCs differentiated toward an endothelial phenotype improves both survival and neovascularization of the transplanted fat lipoaspirate, which might provide benefits and represents a promising strategy for clinical application in autologous fat transplantation.

International audience; As part of normal development most eukaryotic organisms, ranging from insects and mammals to plants, display variations in nuclear ploidy levels resulting from somatic endopolyploidy. Endoreduplication is the major source of endopolyploidy in higher plants. Endoreduplication is a remarkable characteristic of the fleshy pericarp tissue of developing tomato fruits, where it establishes a highly integrated cellular system that acts as a morphogenetic factor supporting cell growth. However, the functional significance of endoreduplication is not fully understood. Although endoreduplication is thought to increase metabolic activity due to a global increase in transcription, the issue of gene-specific ploidy-regulated transcription remains open. To investigate the influence of endoreduplication on transcription in tomato fruit, we tested the feasibility of a RNA sequencing (RNA-Seq) approach using total nuclear RNA extracted from purified populations of flow cytometry-sorted nuclei based on their DNA content. Here we show that cell-based approaches to the study of RNA-Seq profiles need to take into account the putative global shift in expression between samples for correct analysis and interpretation of the data. From ploidy-specific expression profiles we found that the activity of cells inside the pericarp is related both to the ploidy level and their tissue location.

In breast reconstructive procedures, adipose-derived stem cells (ASCs) that are present in clinical fat grafting isolates are considered to play the main role in improving wound healing. In patients following chemotherapy for breast cancer, poor soft tissue wound healing is a major problem. However, it is unclear if tamoxifen (TAM) as the most widely used hormonal therapeutic agent for breast cancer treatment, affects the ASCs and ultimately wound healing. This study evaluated whether TAM exposure to in vitro human ASCs modulate cellular functions. Human ASCs were isolated and treated with TAM at various concentrations. The effects of TAM on cell cycle, cell viability and proliferation rates of ASCs were examined by growth curves, MTT assay and BrdU incorporation, respectively. Annexin V and JC-1 Mitochondrial Membrane Potential assays were used to analyze ASC apoptosis rates. ASCs were cultured in derivative-specific differentiation media with or without TAM (5 uM) for 3 weeks. Adipogenic and osteogenic differentiation levels were measured by quantitative RT-PCR and histological staining. TAM has cytotoxic effects on human ASCs through apoptosis and inhibition of proliferation in dose- and time-dependent manners. TAM treatment significantly down-regulates the capacity of ASCs for adipogenic and osteogenic differentiation (p

Objectives: The goal of our study is to investigate the effect of adipose derived stem cells (ASCs), endothelial differentiated (ED) ASCs, and various conditioned medium (CM) on wound healing in a diabetic swine model. Methods: Diabetes was induced in 4 male Yorkshire pigs via intravenous injection of 150 mg/kg Streptozotocin. Pig ASCs were harvested and cultured in either M199 or EGM-2 medium with 30 mm glucose. Fourteen circular dorsal wounds (5 cm diameter; 5 mm depth) were created. Duplicate wounds were treated with one of 4 cell-based therapies by injection of 5 million (M) ASCs, 10M ASCs, 5M ED ASCs, or 10M ED ASCs on day 0 and half the initial dose on day 15. Wounds assigned to the topical CM therapy were covered with 2 cc of either serum free M199 primed by ASCs or Human Umbilical Vein Endothelial Cells every 3 days. One set of wound was designated as control. Wounds were assessed at day 0, 10, 15, 20 and 28. Animals were sacrificed on day 28. Image J software was used to evaluate percent wound healing. Results: On day 10, 15, 20 and 28 there was a significant increase in percent wound closure rates in both cell-based treatments when compared to control (pFigure 1 ). All treatments displayed significant increase in wound closure rates on day 28 when compared to control. Histology revealed a decrease in dermal inflammatory cells and improved restoration of normal skin architecture in therapeutic treatment groups when compared to untreated controls. Conclusions: Cellular therapy with ASCs, ED ASCs and topical CM accelerate diabetic wound healing in the swine model. Cellular therapy may provide the greatest benefit, as cells may continuously supply growth factors and adapt to the evolving microcellular environment as the wound progresses through the various phases of healing

Chronic wounds are a common surgical problem exacerbated by diabetes and ischemia. Although adipose-derived stem cells (ASCs) have shown promise as a wound healing therapy, their function and proliferation are hindered in diabetes. This study examines the ability of the human umbilical vein endothelial cell (HUVEC) secretome to reverse the deleterious effects of high glucose concentrations on ASCs through priming, thereby enhancing their ability to participate in angiogenesis and wound healing.Institutional review board-approved human ASCs were cultured in M199 medium with or without glucose (30 mmol/L). HUVEC were grown in 30 mmol/L glucose-containing M199 medium; the resulting conditioned medium (HUVEC-CM) was collected every 3 days and used to prime ASCs. An aliquot of HUVEC-CM was heated (85°C for 30 minutes) to produce thermal denaturation of protein. Viability, proliferation, and endothelial differentiation were measured by MTT assays, growth curves, and quantitative polymerase chain reaction, respectively. A Matrigel assay was used to assess the ability of primed ASCs to participate in capillary-like tube formation. An Institutional Animal Care and Use Committee-approved in vivo murine model of diabetic and ischemic hindlimbs was used to evaluate the angiogenic potential of primed stem cells. Human ASCs were cultured with either control M199 or HUVEC-CM. Mice were randomized to a control group, an unprimed ASC group, or a HUVEC-primed ASC group. Cellular therapies were injected into the ischemic muscle. Thirty days later, slides were made. Microvessels were counted by three blinded observers.MTT assays revealed that HUVEC-priming induced a 1.5 times increase in cell viability over diabetic controls. This promoting effect was lost with heated HUVEC-CM (P .001), indicating that the active molecules are of protein origin. After 9 days, ASCs cultured in 30 mmol/L glucose solution showed a 14% reduction in growth from nondiabetic controls (P = .013) and exhibited atrophic morphology. Conversely, diabetic HUVEC-primed stem cells demonstrated a nearly four-fold increase in proliferation (P .05) and took on a fusiform, endothelial-like phenotype. Polymerase chain reaction demonstrated enhanced expression of CD31 messenger RNA by 4.7-fold after 14 days in the HUVEC-primed group, and endothelial nitric oxide synthase messenger RNA messenger RNA was increased 20.1-fold from controls. Unlike unprimed controls, HUVEC-primed ASCs readily formed capillary-like tube networks on Matrigel. Diabetic mice that were injected with HUVEC-primed ASCs demonstrated greater vessel density than both controls (2.1-fold) and unprimed stem cell treatments (P .001).HUVECs secrete protein factors that significantly increase proliferation and endothelial differentiation of ASCs under diabetic conditions. Injection of ischemic hindlimbs in diabetic mice with HUVEC-primed ASCs leads to enhanced angiogenesis.

Background: Diabetes type 1 and 2 cause hyperglycemia and result in endothelial dysfunction with endothelial vessel and poor wound healing. Adipose-derived stem cells (ASCs), progenitor cells in wound healing, show decreased function under hyperglycemic conditions in vitro and in vivo . We hypothesized that exposing ASCs in the presence of high glucose with the human umbilical vein endothelial cell (HUVEC) secretome will reverse the deleterious effects of glucose on ASCs and subsequently enhance angiogenesis and wound healing. Methods: Human umbilical vein endothelial cells (HUVEC) were treated with glucose (30mM) and the conditioned media (CM) were collected every 3 days. ASCs were then co-cultured with EC/CM for 2 weeks. To produce thermal denaturation of protein, EC/CM was heated at 95 0 C for 30 mins. Cell activity, proliferation, and endothelial-like properties of ASCs were determined by MTT assays, growth curves, and real-time RT-PCR, respectively. EC/CM treated ASC were injected into a normal or diabetic murine left thigh muscle at three different points with hindlimb ischemia. After 4 weeks injection, animals were sacrificed. H & E and double immunostaining for CD31 and anti-human nuclei were used to determine if the ASCs primed with EC/CM underwent neovascularization. Results: In fact, ASCs increased in proliferation when co-cultured with HUVEC/CM (1.4 fold) when compared with controls. This promoting effect was lost in heated HUVEC/CM, indicating that the active molecules are of protein origin. After 10 days stimulated with EC/CM an increase in mRNA expression levels of EC markers were also observed in high glucose (30mM) EC/CM environment including CD31 (2-fold), vWF (1.1-fold), and eNOS (3.2-fold) when compared to ASCs cultured in M199. H & E and immunohistochemical staining results showed elevated vessel density and CD31 + cell levels in HUVEC-primed ASC injection sites of diabetic mice when compared with the control animals. Conclusions: HUVEC secrete protein factors that increase proliferation and endothelial differentiation of ASCs under diabetic conditions. Injection of ischemic hindlimbs in diabetic mice with HUVEC-primed ASCs leads to improved angiogenesis.

The legume genus Aeschynomene is notable in the ability of certain semiaquatic species to develop nitrogen-fixing stem nodules. These species are distributed in two clades. In the first clade, all the species are characterized by the use of a unique Nod-independent symbiotic process. In the second clade, the species use a Nod-dependent symbiotic process and some of them display a profuse stem nodulation as exemplified in the African Aeschynomene afraspera. To facilitate the molecular analysis of the symbiotic characteristics of such legumes, we took an integrated molecular and cytogenetic approach to track occurrences of polyploidy events and to analyze their impact on the evolution of the African species of Aeschynomene. Our results revealed two rounds of polyploidy: a paleopolyploid event predating the African group and two neopolyploid speciations, along with significant chromosomal variations. Hence, we found that A. afraspera (8x) has inherited the contrasted genomic properties and the stem-nodulation habit of its parental lineages (4x). This study reveals a comprehensive picture of African Aeschynomene diversification. It notably evidences a history that is distinct from the diploid Nod-independent clade, providing clues for the identification of the specific determinants of the Nod-dependent and Nod-independent symbiotic processes, and for comparative analysis of stem nodulation.

Background: Topical lidocaine is a common form of anesthesia for a wealth of procedures across a large number of disciplines, including laser treatments. Preparations can be purchased over the counter with no prescription necessary. It is considered a safer and more acceptable form of anesthetic than hypodermic injections; however, there have been reports of fatalities following its application. Above certain serum lidocaine concentrations, patients may experience effects of toxicity such as lightheadedness and paraesthesia; these effects can progress to seizures and cardiorespiratory depression, which can ultimately lead to death. The active metabolite of lidocaine, monoethylglycinexylidide (MEGX), can be almost as potent as lidocaine in terms of toxicity. Objectives: The authors examine the levels of both lidocaine and MEGX in blood serum after application of topical lidocaine. Methods: Twenty-five healthy volunteers were assigned to one of four groups (A, B, C, D). Group A had 2.5 g of 4% lidocaine topical anesthetic cream applied to the face for one hour without occlusion, Group B had 5 g applied to the face for one half-hour without occlusion, Group C had 5 g applied to the face for one hour without occlusion, and Group D had 5 g applied to the face for one hour with occlusion. To evaluate serum concentrations, blood was drawn every 30 minutes for four hours. Results: Group D showed the highest serum levels of lidocaine and MEGX, a three-fold increase compared with group C, which received the same dose (5g topical 4% lidocaine) but without occlusion. In group D, peak serum levels occurred at 90 minutes for serum lidocaine, which was also the fastest of the four groups. Serum MEGX levels peaked much later than serum lidocaine levels, at 210 minutes. Individual serum levels did not exceed 0.6 µg/mL. Across the groups, there was significant interindividual variation in both lidocaine and MEGX serum levels ( P = .061). Applications of 5 g of 4% lidocaine resulted in higher serum concentration of both lidocaine and MEGX. When comparing group A to group C, doubling the dose of 4% lidocaine from 2.5 g to 5 g resulted in double the serum levels of MEGX and a 50% increase in the serum lidocaine levels ( P = .021). When comparing groups C and D, the addition of an occlusive dressing resulted in a tripling of the serum lidocaine levels and a doubling of the serum MEGX levels, both of which were statistically significant ( P < .001). When comparing all four groups, there were significant differences between the combined serum concentrations of lidocaine and MEGX ( P < .001). Conclusions: Topical lidocaine preparations are increasingly being employed to provide a patient-friendly form of noninvasive analgesia for a multitude of procedures. Some preparations are available over the counter for unsupervised patient application. Our study has demonstrated significant interindividual variability for a given dose, especially when occlusion is applied. There have been fatalities resulting from topical lidocaine application, and our study suggests that this is the result of the unpredictability of lidocaine metabolism between individuals. Therefore, we recommend that caution be exercised with topical lidocaine preparations, in particular when applied in conjunction with occlusive dressings.

Background: This is a report of the histopathological evaluation of the acute damage profile in human skin following treatment with two novel short-pulsed fractional carbon dioxide resurfacing devices used independently and in combination in vivo. Methods: The panni of eight abdominoplasty patients were treated with either the Active FX, the Deep FX (Lumenis Ltd., Yokneum, Israel), or a combination of the two (Total FX) prior to the start of the excisional surgical procedure. Multiple combinations of energies, pulse widths, and densities were evaluated for each device. After surgical removal (two to five hours), each pannus was immediately biopsied and samples were processed for histopathological evaluation. Results: The Active FX system resulted in extensive epidermal injury with wide shallow ablation craters that, at higher fluences, extended through the basement membrane of the epidermis into the papillary dermis. The Deep FX fractional treatment caused deep microcolumns of ablation penetrating up to 3 to 4 mm from the epidermal surface into the deep reticular dermis with a variable rim of coagulated collagen surrounding each ablation column. Conclusions: The in vivo histopathological evaluation of these devices furthers our understanding of the fundamental laser/tissue interaction following treatment with each device independently and in combination.

Background: Both erbium (Er:YAG) and carbon dioxide (CO2) devices are commonly-used, efficient laser systems for aggressive skin resurfacing procedures. The devices each have different adjustable parameters (density, spot size, number of pulses, pattern, etc) and utilize variable energy capabilities to tailor individual treatments depending on the skin pathology and goals of treatment. Overall, the consensus has been that multiple-pass erbium treatments needed for efficacious wrinkle reduction had similar downtime and comorbidity to the traditional CO2 treatments. Unfortunately, there were limited data comparing the histological differences and changes throughout the wound-healing process over time between the two treatment methods. Objectives: The authors compare the difference in injury following treatment with five novel fractional ablative laser systems in vivo. Differences in damage pattern, treatment depth, and degree of surrounding cellular injury following treatment with each device at common clinical settings are evaluated in a side-by-side histopathologic comparison. Methods: Prior to planned excisional surgery, the panni of 20 abdominoplasty patients were treated with five novel ablative fractional carbon dioxide or Er:YAG laser systems at various clinical parameters, in accordance with the manufacturers’ treatment guidelines. After tissue removal two to four hours later, the skin was biopsied and processed for histopathologic evaluation. Specimens were stained with hematoxylin and eosin, along with a terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling (TUNEL) assay, to highlight the degree of irreversible cellular injury. Results: The acute damage profile differed between the fractional Er:YAG and CO2 devices with respect to depth of penetration and extension of coagulation surrounding the microcolumns. The damage pattern was dependent on the parameters set with each device (eg, fluence, pulses, density, pulse width). The TUNEL-stained sections demonstrated more collateral cellular injury surrounding the ablated columns with the CO2 devices than with the Er:YAG systems. Conclusions: Following treatment with the fractional Er:YAG and CO2 devices, deep tissue injury with various coagulative and ablative properties was observed, and it was confirmed that carbon dioxide and erbium devices result in different patterns of injury. As such, each may be better suited for different clinical situations. It is important for practitioners to understand the limitations of a specific device, as well as the tissue injury following a given treatment pattern or protocol, to appropriately tailor their treatment algorithm for a given patient. This extensive histopathologic evaluation of the acute characterization of injury across devices is helpful in clarifying the differences/similarities in laser-tissue interaction following treatment in an in vivo human model.

A study was undertaken to survey current practice patterns concerning primary breast augmentation. Members of the American Society for Aesthetic Plastic Surgery (ASAPS) were electronically surveyed concerning issues such as incision location, implant size and type, and complications, as well as information about the surgeons, their practices, and where procedures are performed. The survey response rate was 30%. Plastic surgeons from the South and Southwest made up 40% of respondents. Forty-six percent of respondents had more than 20 years of experience in practice. Forty-three percent of primary breast augmentations were performed in outpatient surgery centers. An anesthesiologist was in attendance in 60% of cases. The average operative time--indicated in 80% of responses--ranged from 45 to 90 minutes. Thirty-three percent of responding plastic surgeons used the base diameter to determine implant size and respondents most commonly used a smooth saline implant placed through an inframammary incision in a submuscular pocket. The most frequently reported complication was nipple sensation changes. Although the reintroduction of silicone gel implants was accompanied by expectations of a sharp increase in their use, this survey revealed that among ASAPS members, saline implants currently are used more often than silicone gel implants. However, both saline and silicone gel implants are used frequently, safely, and reliably. This survey represents a snapshot of current practice and future trends in primary breast augmentation will require additional assessment, although increased use of silicone gel breast prostheses over time is expected.

Background: Injectable hyaluronic acid fillers (HAFs) and laser/light procedures have become increasingly popular for noninvasive facial rejuvenation in many cosmetic practices. However, the effect of laser/light treatments on HAFs is unknown. Objective: Our objective was to examine the effect of laser/light treatments on HAFs in a porcine model. Methods: The abdomens of 6 Yorkshire pigs were injected with 3 different HAFs: Restylane (Medicis, Scottsdale, AZ), Perlane (Medicis), and Juvederm (Allergan, Santa Barbara, CA). Two weeks after injection, the injection sites were treated with 1 of 7 common laser/light ablative or nonablative devices. Following laser treatment, 8-mm punch biopsies were collected from the treated tissue and fixed for histopathologic evaluation. Sections were stained with hematoxylin–eosin and alcian blue stains for identification of the preinjected HAF. Results: The filler was identified in different areas of the dermis in different sections. The Sciton intense pulsed 560 nm filter (Sciton, Palo Alto, CA), the Sciton Nd: YAG, Lux1540 (Palomar Medical Technologies, Burlington, MA), or ActiveFX (Lumenis, Yokneum, Israel) treatments showed no sign of interaction with superficial or deep dermal filler. No evidence of morphologic changes to the filler or the surrounding tissues was observed. Obvious interaction between the HAFs and the laser injury was demonstrated in sections treated with the deep ablative systems fractional erbium 2940 (Profractional; Sciton) and DeepFX CO2, (Lumenis). However, no uncharacteristic tissue injury or morphologic change in the filler was appreciated in any of the preinjected specimens. Conclusions: Injected HAFs were unaffected by the nonablative laser/light and superficial ablative treatments. The more aggressive deeper laser treatments demonstrated laser/filler interaction and may have a clinical effect on the longevity of the filler and/or efficacy of laser treatments. Novel ablative fractional lasers have the capability of deep dermal penetration, and this should be taken into consideration when planning to use them in combination with soft tissue fillers for noninvasive facial rejuvenation.

Rationale and Objectives Thinned perforator flaps have been widely used in plastic surgery for greater survivability and decreased morbidity. However, quantitative analysis of three-dimensional (3D) blood flow direction and location has not been examined yet. Such information will benefit and guide the surgical thinning and dissection process. Toward this goal, this study was performed for 3D vascular tree reconstruction with the incorporation of temporal contrast-agent propagation information (three spatial dimensions plus one temporal dimension; ie, 4D). Materials and Methods A novel computational framework by adopting a moving grid deformation method is presented. To take advantage of temporal information of the bolus propagating, a sequential segmentation procedure is proposed. Moreover, the temporal evolution of the vascular tree (4D vascular tree) is reconstructed during the procedure. Results Eight anterolateral thigh perforator flaps from eight cadavers were used for this study. The age range is 60–80 years old and the gender includes four males and four females. The 3D nature of the vascular structure and 4D vascular tree evolving process are showed in comparison with maximum intensity projection images. Conclusion The proposed computational framework demonstrates effectiveness in the modeling of 4D vascular tree. Furthermore, it reveals the ability to detect small vessel tree structures that are beyond the limit of image resolution.

The author stresses that to date, the effects of mesotherapy have not been scientifically evaluated. Currently, there is no standardization of dosage and no protocol or treatment algorithm to enable prediction of how much tissue or fat will be "dissolved" with a specific solution in a defined quantity, and injected at a specified subcutaneous tissue depth.

The cell division cycle involves nuclear and cytoplasmic events, namely organelle multiplication and distribution between the daughter cells. Until now, plastid and plant cell division have been considered as independent processes because they can be uncoupled. Here, down-regulation of AtCDT1a and AtCDT1b, members of the prereplication complex, is shown to alter both nuclear DNA replication and plastid division in Arabidopsis thaliana . These data constitute molecular evidence for relationships between the cell-cycle and plastid division. Moreover, the severe developmental defects observed in AtCDT1-RNA interference (RNAi) plants underline the importance of coordinated cell and organelle division for plant growth and morphogenesis.

The anaphase-promoting complex (APC), a multisubunit E3 ubiquitin ligase, is an essential regulator of the cell cycle from metaphase until S phase in yeast and metazoans. APC mediates degradation o ...

Biologic scaffolds composed of naturally occurring extracellular matrix (ECM) can provide a microenvironmental niche that alters the default healing response toward a constructive and functional outcome. The present study showed similarities in the remodeling characteristics of xenogeneic ECM scaffolds when used as a surgical treatment for volumetric muscle loss in both a preclinical rodent model and five male patients. Porcine urinary bladder ECM scaffold implantation was associated with perivascular stem cell mobilization and accumulation within the site of injury, and de novo formation of skeletal muscle cells. The ECM-mediated constructive remodeling was associated with stimulus-responsive skeletal muscle in rodents and functional improvement in three of the five human patients.

Background: Chronic wound healing problems can pose a significant clinical challenge. Transdermal delivery of adipose-derived stem cells (ADSC) may be a possible solution to healing these recalcitrant, debilitating wounds. Pretreatment of the skin with a fractionated laser has already been shown to assist transdermal drug delivery both in vitro and in vivo and may be an ideal approach to facilitating delivery of ADSC to the target tissue. Objectives: The authors investigate in a porcine model whether ADSC can be delivered transdermally following pretreatment with a fractional laser. Methods: After ethics approval was obtained, the abdomens of 2 adult female domestic pigs were pretreated with an erbium:YAG fractionated ablative laser. Following laser treatment, 20 × 106 bromodeoxyuridine (BrdU)–labeled ADSC were applied topically to the first animal for 4 hours. The same number of BrdU-labeled ADSC was applied to the second animal for 48 hours. The animals were euthanized at the end of their respective treatment periods, and the BrdU-labeled ADSC were counted after tissue harvest. Results: At 4 hours, an average of 2.40 × 106 cells, or 12.0% of the total cells applied, were found in the tissue. At 48 hours, an average of 1.1 × 106 cells, or 5.5% of the total cells applied, were seen. Conclusions: This pilot study demonstrates that ADSC can be delivered transdermally through skin that has been pretreated with a laser. Potential future applications of this approach might include wound-healing or aesthetic indications. Further studies need to be conducted to determine the optimal number of ADSC to use in this approach, the best methods of application, and the effect of transdermally delivered ADSC on wound healing.

Background: Topical anesthetics are commonly applied for a variety of indications. Several lidocaine-containing topical anesthetics are available for purchase over the counter (OTC). Recently, the authors’ group has shown that there is great interindividual discrepancy in the manner in which lidocaine is absorbed and metabolized for even a single OTC product. Objectives: The authors compare five commonly-available lidocaine preparations and their levels of absorption when applied to the face. Three of the preparations are available OTC; two require prescriptions and were compounded in a pharmacy. Methods: Twenty-five subjects enrolled in this Institutional Review Board–approved study were randomly assigned to one of five groups. The five topical anesthetics were LMX-4 (4% lidocaine; Biopelle/Ferndale Laboratories, Ferndale, Michigan), Topicaine (4% lidocaine; Ebsa Laboratories, Jupiter, Florida), 2.5% lidocaine/2.5% prilocaine (generic EMLA preparation; High Tech Pharmaceuticals, Amityville, New York), LET (4% lidocaine, 1:2000 epinephrine, and 0.5% tetracaine), and BLT (20% benzoncaine, 6% lidocaine, and 4% tetracaine). After a patch test for adverse reactions, the topical anesthetic was applied to each patient’s face and neck and covered with an occlusive dressing for 60 minutes. Blood was drawn at 90, 120, 150, 240, and 480 minutes to measure serum levels of lidocaine and monoethylglycinexylidide (MEGX). Results: The average age of the 17 women and eight men included in the study was 26 years (range, 22-62 years), and the average weight was 70.9 kg (range, 46.4-96.4 kg). The OTC preparations had the highest serum lidocaine and MEGX levels. Topicaine had the greatest serum levels of individual lidocaine absorption (0.808 µg/mL), followed by generic EMLA (0.72 µg/mL), LMX-4 (0.44 µg/mL), BLT (0.17 µg/mL), and LET (0.13 µg/mL). On average, Topicaine had the highest serum lidocaine and MEGX levels: 0.438 µg/mL and 0.0678 µg/mL, respectively. There were significant interindividual differences between the serum levels of MEGX and lidocaine in all groups except LET ( P < .0001). There were significant differences between the 4% lidocaine-containing preparations ( P = .0439); the 2.5% preparation had a greater absorption than the 4% lidocaine-containing preparation and the 6% lidocaine preparation ( P = .0016). There were three adverse reactions in patients who received OTC preparations, one of which resulted in postinflammatory hyperpigmentation. Conclusions: This study demonstrates that although topical anesthetics are considered safe, some individuals have unpredictably high absorption levels. This study also demonstrates that the concentration of lidocaine, the formulation of the drug, and the individual patient all have significant effects on serum levels of lidocaine. The authors recommend that even OTC topical anesthetics be used under the supervision of a healthcare professional to avoid adverse toxic effects and, in rare cases, death. Level of Evidence: 2 ![Graphic][1] [1]: /embed/inline-graphic-1.gif

Individuals with elevated levels of plasma cholesterol and triglyceride may be at higher risk for coronary artery disease than those with isolated elevations of either cholesterol or triglyceride. Sequence variation in the A-I/C-III/A-IV gene cluster has been implicated in the etiology of some disorders associated with premature atherosclerosis and/or hypertriglyceridemias with or without elevations of cholesterol. This led to the hypothesis that allelic variation at this gene locus alters plasma lipid transport and affects susceptibility for atherosclerosis. The study population, from the Atherosclerosis Risk in Communities (ARIC) Study, consisted of 50 normolipidemic individuals, 48 subjects with elevated plasma cholesterol, 47 subjects with elevated plasma triglyceride, and 123 subjects with both elevated plasma cholesterol and triglyceride who were used to evaluate associations between an Xmn I polymorphic site 2.5 kilobase pairs (kbp) upstream of the structural gene for apolipoprotein (apo) A-I, intimal-medial thickening of the extracranial carotid arteries, and several plasma lipid factors. The relative allele frequencies of the 8.3-kbp allele and the 6.6-kbp allele were .86 and .14, respectively, in the entire study population and did not differ among the lipid phenotypes. In the group with elevated plasma cholesterol and triglyceride, subjects possessing the 6.6-kbp allele exhibited a greater carotid artery intimal-medial thickness (P = .034) and higher plasma levels of apoA-I, high-density lipoprotein (HDL) cholesterol, and HDL3 cholesterol (P < .02) than subjects homozygous for the 8.3-kbp allele. In contrast, subjects with the 6.6-kbp allele displayed lower mean ratios of apolipoproteins C-II to C-III, C-II to A-IV and E to A-IV in plasma (P < .05) and a lower mean ratio of apolipoprotein C-II to C-III in the triglyceride-rich lipoproteins (P = .026). Sequence variation in or near the genes encoding apolipoproteins A-I, C-III, and A-IV may therefore identify a group of hypercholesterolemic-hypertriglyceridemic persons who are at higher risk for atherosclerosis than others with the same lipoprotein phenotype.

To gain insight into metabolic determinants of high density lipoproteins (HDL) containing apolipoproteins A-I and A-I1 (LpA-I/A-11) and those containing A-I, but devoid of A-I1 (LpA-I), the plasma concentration of LpA-I and LpA-I/A-I1 wi- thin the HDLz and HDL3 density spectrum was measured in 14 normolipidemic male subjects on a standardized diet. Apolipoprotein plasma concentrations of HDL subspecies were compared with the magnitude of postprandial lipemia, activities of lipoprotein lipase and hepatic lipase in postheparin plasma, plasma 1ecithin:cholesterol acyltransferase (LCAT) activity, and cholesteryl ester transfer protein (CETP) mass. Plasma levels of LpA-I/A-I1 were 2.5 times higher than levels of LpA-I (123 + 20 vs. 48.3 f 22.1 mg protein/dl) and the partition of LpA-I and LpA-I/A-I1 between HDLz and HDL3 differed in that the proportion of LpA-I associated with HDLZ was greater than that of LpA-I/A-I1 (23 f 19 vs. 6 6%, P < 0.002). With increas- ing levels of HDLZ, the proportion of LpA-I in HDLZ increased (P < 0.002). Furthermore, levels of LpA-I and LpA-I/A-I1 were strongly correlated within the HDLZ, but not within the HDLS density region. Plasma levels of LpA-I, but not LpA-I/A-11, were inversely correlated with the magnitude of postprandial lipemia. However, activities of lipoprotein lipase and hepatic lipase tended to show stronger associations with the partition of LpA- I/A-I1 between HDLz and HDL3 than with that of LpA-I. Wi- thin the HDL,, but not the HDLZ density spectrum, LpA-I/A-I1 exhibited a positive association with plasma LCAT activity, while LpA-I displayed an inverse association with plasma CETP mass. I These results are consistent with differences in sub- strate properties of LpA-I and LpA-I/A-I1 for lipoprotein- modifying enzymes and imply different, but overlapping meta- bolic pathways of LpA-I and LpA-I/A-11.-Mowri, H-O., J. R. Patsch, A. Ritsch, B. Foger, S. Brawn, and W. Patsch. High density lipoproteins with differing apolipoproteins: relationships to postprandial lipemia, cholesteryl ester transfer protein, and activities of lipoprotein lipase, hepatic lipase, and leci- thin:cholesterol acyltransferase. J. Lipid Res. 1994. 35: 291-300.

Background: The mechanism(s) responsible for breast capsular contracture (CC) remain unknown, but inflammatory pathways play a role. Various molecules have been attached to implant shells in the hope of modifying or preventing CC. The intrinsic antibacterial and antifungal activities of chitosan and related oligochitosan molecules lend themselves well to the study of the infectious hypothesis; chitosan's ability to bind to growth factors, its hemostatic action, and its ability to activate macrophages, cause cytokine stimulation, and increase the production of transforming growth factor (TGF) β1 allow study of the hypertrophic scar hypothesis. Objective: The authors perform a comprehensive evaluation, in a rabbit model, of the relationship between CC and histological, microbiological, and immunological characteristics in the presence of a chitooligosaccharide (COS) mixture and a low molecular weight chitosan (LMWC). Methods: Eleven adult New Zealand rabbits were each implanted with three silicone implants: a control implant, one impregnated with COS, and one impregnated with LMWC. At four-week sacrifice, microdialysates were obtained in the capsule-implant interfaces for tumor necrosis factor alpha (TNF-α) and interleukin-8 (IL-8) level assessment. Histological and microbiological analyses were performed. Results: Baker grade III/IV contractures were observed in the LMWC group, with thick capsules, dense connective tissue, and decreased IL-8 levels (p < .05) compared to control and COS groups. Capsule tissue bacterial types and microdialysate TNF-αlevels were similar among all groups. Conclusions: Chitosan-associated silicone implantation in a rabbit model resulted in Baker grade III/IV CC. This preclinical study may provide a model to test various mechanistic hypotheses of breast capsule formation and subsequent CC.

Background: The etiology and ideal clinical treatment of capsular contracture (CC) remain unresolved. Bacteria, especially coagulase-negative staphylococci, have been previously shown to accelerate the onset of CC. The role of fibrin in capsule formation has also been controversial. Objective: The authors investigate whether fibrin and coagulase-negative staphylococci (CoNS) modulate the histological, microbiological, and clinical outcomes of breast implant capsule formation in a rabbit model and evaluate contamination during the surgical procedure. Methods: Thirty-one New Zealand white female rabbits were each implanted with one tissue expander and two breast implants. The rabbits received (1) untreated implants and expanders (control; n = 10), (2) two implants sprayed with 2 mL of fibrin and one expander sprayed with 0.5 mL of fibrin (fibrin; n = 11), or (3) two implants inoculated with 100 µL of a CoNS suspension (108CFU/mL—0.5 density on the McFarland scale) and one expander inoculated with a CoNS suspension of 2.5 × 107 CFU/mL (CoNS; n = 10). Pressure/volume curves and histological and microbiological evaluations were performed. Operating room air samples and contact skin samples were collected for microbiological evaluation. The rabbits were euthanized at four weeks. Results: In the fibrin group, significantly decreased intracapsular pressures, thinner capsules, loose/dense (

Background: The root cause of capsular contracture (CC) associated with breast implants is unknown. Recent evidence points to the possible role of fibrin and bacteria in CC formation. Objectives: The authors sought to determine whether fibrin, thrombin, and blood modulated the histological and microbiological outcomes of breast implant capsule formation in a rabbit model. Methods: The authors carried out a case-control study to assess the influence of fibrin, thrombin, and blood on capsule wound healing in a rabbit model. Eighteen New Zealand white rabbits received four tissue expanders. One expander acted as a control, whereas the other expander pockets received one of the following: fibrin glue, rabbit blood, or thrombin sealant. Intracapsular pressure/volume curves were compared among the groups, and histological and microbiological evaluations were performed (capsules, tissue expanders, rabbit skin, and air). The rabbits were euthanized at two or four weeks. Results: At four weeks, the fibrin and thrombin expanders demonstrated significantly decreased intracapsular pressure compared to the control group. In the control and fibrin groups, mixed inflammation correlated with decreased intracapsular pressure, whereas mononuclear inflammation correlated with increased intracapsular pressure. The predominant isolate in the capsules, tissue expanders, and rabbit skin was coagulase-negative staphylococci. For fibrin and thrombin, both cultures that showed an organism other than staphylococci and cultures that were negative were associated with decreased intracapsular pressure, whereas cultures positive for staphylococci were associated with increased intracapsular pressure. Conclusions: Fibrin application during breast implantation may reduce rates of CC, but the presence of staphylococci is associated with increased capsule pressure even in the presence of fibrin, so care should be taken to avoid bacterial contamination.

Plasma levels of lipoprotein[a] (Lp[a]) are associated with increased risk of coronary artery disease and show an inverse correlation with apolipoprotein[a] (apo[a]) molecular weight. We determined Lp[a] levels and apo[a] phenotypes in 171 cases with preclinical extracranial carotid atherosclerosis as ascertained by B-mode ultrasound and in 274 control subjects free of carotid atherosclerosis. Lp[a] protein levels measured by enzyme-linked immunosorbent assay ranged from 4 to 361 micrograms/mL in cases and from 2 to 392 micrograms/mL in controls, but median levels of Lp[a] were higher in cases than in controls (51 micrograms/mL versus 33 micrograms/mL, P < .003). In both groups, all 11 apo[a] polymorphs that are resolved by the procedure used were present, resulting in 43 and 39 different apo[a] phenotypes in cases and controls, respectively. An inverse relation between apo[a] polymorph size and Lp[a] level was observed in both cases (r = -0.49, P < .001) and controls (r = -0.34, P < .001). Apo[a] phenotype distributions were similar in cases and controls. However, in 17 phenotypes with three or more subjects per group, the difference of mean Lp[a] concentrations between cases and controls was 32 +/- 36 micrograms/mL (mean +/- SD). Thus, the higher Lp[a] levels in cases were not associated with a greater prevalence of small apo[a] polymorphs. Stepwise logistic regression analyses of known risk factors for coronary heart disease showed that plasma Lp[a] concentration was an independent predictor of case-control status, while Lp[a] phenotype was not, irrespective of the presence or absence of Lp[a] concentration in the model.(ABSTRACT TRUNCATED AT 250 WORDS)

The distributions of plasma lipids, lipoproteins, and apoproteins of 14,524 female and male black and white participants 45 to 64 years old in the Atherosclerosis Risk in Communities (ARIC) Study are presented. All specimens were analyzed at a central laboratory. Mean total cholesterol levels increased with increasing age across all ages from 204 to 229 mg/dL (12%) in women and from 208 to 213 mg/dL (2%) in men. Triglyceride levels increased with age in women, remained stable in men, and were higher in whites than blacks. High-density lipoprotein (HDL) cholesterol levels were higher in black and white women (range, 57 to 59 mg/dL) compared with black men (49 to 52 mg/dL) or white men (42 to 43 mg/dL). Cholesterol associated with HDL was distributed in a relatively constant proportion between HDL3 (70% to 76%) and HDL2 (24% to 30%) for all race/sex groups. Low-density lipoprotein (LDL) cholesterol levels increased with age in black (14.7%) and white (17.1%) women and in black (4.4%) and white (3.7%) men; more than 50% of all participants had LDL cholesterol levels > 130 mg/dL. Apoprotein A-I and B levels followed the same trends as HDL cholesterol and LDL cholesterol levels, respectively. Lipoprotein(a) [Lp(a)] levels were twice as high in blacks as in whites, and women's Lp(a) levels were higher than men's Lp(a) levels for each race. Menopause was associated with elevated total cholesterol, LDL cholesterol, apoprotein B, and Lp(a) levels, and hormone replacement medication use in postmenopausal subjects was associated with higher HDL cholesterol, triglyceride, and apoprotein A-I levels and lower LDL cholesterol, apoprotein B, and Lp(a) levels.