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Biomedical subjects

John C Lee

Publications and source records attributed to John C Lee.

At least 19 recordsLinked to original sources
PubMed2006

Bone morphogenetic protein-4 enhances vascular endothelial growth factor secretion by human retinal pigment epithelial cells.

Retinal pigment epithelial (RPE) cells secrete vascular endothelial growth factor (VEGF), a cytokine known to promote angiogenesis. Results from RNase protection assays (RPAs) show that RPE from non-diabetic human donors and from adult retinal pigment epithelium-19 (ARPE-19) cells expressed significant bone morphogenetic protein-4 (BMP-4) message. In addition, ARPE-19 cells cultured in high glucose (25 mM), compared to those in physiological glucose (5.5 mM) released significantly more BMP-4 into the conditioned media (CM). However, the effect of BMP-4 on the release of VEGF by ARPE-19 cells has not been studied. Accordingly, ARPE-19 cells were treated with BMP-4 to determine VEGF secretion. BMP-4 and VEGF levels in the CM and cell lysates were measured by enzyme-linked immunosorbent assay (ELISA). Cells treated with exogenous BMP-4 had higher VEGF in the CM and this treatment effect was dose- and time-dependent, while cell lysates had low levels of VEGF. Addition of cycloheximide (CHX) or actinomycin-D (ACT) significantly reduced VEGF secretion from cells treated with BMP-4, suggesting that the BMP-4-induced secretion of VEGF requires new RNA and protein synthesis. Our results suggest that BMP-4 may play a role in the regulation of ocular angiogenesis associated with diabetic retinopathy (DR) by stimulating VEGF release from RPE cells.

Adult↗
PubMed2006

Differential autonomic nervous system response in obese and anorexic chickens (Gallus gallus).

Effect of reserpine on body weight (BW), feed intake (FI), brain and plasma catecholamine and indoleamine concentrations in high- (HWS) and low- (LWS) weight selected lines of chickens was investigated. Chicks from each line were assigned to three treatment groups and injected intraperitoneally with 0, 1.25, or 2.50 mg/kg of reserpine at hatch, and again at 5 weeks-of-age. Chick BW and FI were determined weekly. At 7 weeks-of-age, 12 males and females from each group were sacrificed for neurotransmitter analysis. In the HWS line there was a dose-dependent decrease in BW through 7 weeks-of-age, whereas in the LWS line BW decreased only through the first 2 weeks-of-age. In the LWS line, norepinephrine (NE), epinephrine, and 3,4-dihydroxyphenylacetate concentrations decreased in the brain in a linear and quadratic manner in response to reserpine, but not in the HWS line. Both lines showed linear decreases in dopamine levels in response to reserpine; however, serotonin was not affected by reserpine. Chickens in the HWS line had greater plasma NE, and lower 5-hydroxyindoleacetic acid than those in the LWS line. In conclusion, it appears that chickens from the HWS line were more sensitive to the BW reducing effects of reserpine than those from the LWS line, with the latter appearing to have greater sympathetic nervous system activity.

Animal Feed↗
PubMed2006

C-type natriuretic peptide enhances osteogenic protein-1-induced osteoblastic cell differentiation via Smad5 phosphorylation.

In the present study, we examined the hypothesis that the C-type natriuretic peptide (CNP) enhances osteogenic protein-1 (OP-1) action in stimulating osteoblastic cell differentiation in primary cultures of fetal rat calvaria cell (FRC). CNP enhanced synergistically the OP-1-induced Alkaline Phosphatase (AP) activity and mineralized bone nodule formation in a dose- and time-dependent manner. To examine possible mechanism of the synergy between OP-1 and CNP, the expression levels of key BMP receptors and signaling molecules were examined. Western blot analysis showed that BMPR-IB and -II receptor protein expression was not affected by CNP alone, but was stimulated by OP-1 alone. The combination of OP-1 and CNP did not further increase their protein levels. The Runx2 protein expression level was not altered by CNP alone, but was elevated by OP-1 alone, and was slightly reduced by the combination. The Smad5 protein expression level was slightly decreased by CNP alone, but was stimulated by OP-1 alone, and was not further stimulated by the combination. Smad5 phosphorylation was not stimulated by CNP alone, but was stimulated significantly by OP-1 alone. The combination of OP-1 and CNP further stimulated the OP-1-induced Smad5 phosphorylation. Thus, one mechanism of the observed synergy between OP-1 and CNP involves enhancement of the Smad5 phosphorylation.

Alkaline Phosphatase↗
PubMed2006

Damage-control laparotomy.

PURPOSE OF REVIEW: This article summarizes the current state of damage-control laparotomy as practiced in trauma surgery. Since the first description of deliberately abbreviated laparotomy 20 years ago, damage-control laparotomy has been widely applied. The purpose of this review is to discuss current concepts in damage-control laparotomy in trauma and general surgery patients. RECENT FINDINGS: The immediate, essential goals of control of surgical bleeding and containment of gastrointestinal soilage are achieved at a truncated laparotomy. Ongoing resuscitation of the injured patient with severe physiologic derangements is continued in the intensive care unit. Only when the lethal triad of hypothermia, metabolic acidosis, and coagulopathy is corrected does the patient subsequently undergo definitive surgery. Recent studies have better defined the subset of patients that benefit from such an approach. SUMMARY: Application of abbreviated laparotomy has been widely applied in the trauma population. Breaking the pathophysiologic cycle of hypothermia, coagulopathy, and acidosis with this approach has improved survivorship in this critically injured group of patients. The extension of the abbreviated laparotomy concept has also been described in the general surgery population, and raises the possibility of extending this concept to broader surgical fields.

Acidosis↗
PubMed2005

Co-transfection with the osteogenic protein (OP)-1 gene and the insulin-like growth factor (IGF)-I gene enhanced osteoblastic cell differentiation.

Previous studies from this laboratory showed that the action of Osteogenic Protein-1 (OP-1, BMP-7) on osteoblastic cell differentiation could be enhanced by other protein factors, such as Insulin-like Growth Factor (IGF)-I. In the present study, we examined the effects of co-transfection with a combination of the OP-1 and the IGF-I gene on osteoblastic cell differentiation. The results first showed that fetal rat calvaria (FRC) cells transfected with the OP-1 gene under the control of the cytomegalovirus (CMV) promoter showed substantial production of the OP-1 protein. Transfected FRC cells also showed a DNA concentration-dependent increase in alkaline phosphatase (AP) activity, an osteoblastic cell differentiation marker. Von Kossa-positive nodules, a hallmark of bone formation in long-term cultures of bone-derived cells, were also observed in the transfected cells after 26 days in culture, whereas none were observed in control cells. Co-transfection of FRC cells with the combination of the OP-1 and the IGF-I gene resulted in a synergistic stimulation of AP activity. The increase was DNA dose-dependent. The current data show that transfection of OP-1 gene into osteoblastic cells stimulates osteoblastic cell differentiation in vitro. The study further demonstrates the feasibility of employing gene transfer of a second gene in combination with an OP-1 vector to synergistically enhance OP-1 activity.

Alkaline Phosphatase↗
PubMed2005

Cartilage-derived morphogenetic proteins induce osteogenic gene expression in the C2C12 mesenchymal cell line.

Cartilage-derived morphogenetic protein-1, -2, and -3 (CDMP-1, -2, and -3) are members of the bone morphogenetic protein (BMP) family and have been shown to exhibit a variety of biological activities. In the present study, effects of these CDMPs on the temporal and spatial expression of genes in the pluripotent mesenchymal cell line C2C12 were examined. Cells cultured in the presence of CDMPs lost the characteristic elongated shape of myoblasts. At the molecular level, CDMP treatment did not change the mRNA expression of MyoD, aggrecan, Six1, and tendin. Scleraxis mRNA level was reduced by CDMP treatment. CDMP-1 and -3, but not CDMP-2, stimulated expression of osteogenic markers, such as alkaline phosphatase (AP), osteocalcin (OC), BSP, and type I collagen, in a dose- and time-dependent manner. With few exceptions, the three CDMPs changed, with different potencies, the expression profile of different members of the BMP family in a similar temporal pattern. Except at the late phase of treatment, CDMP treatment did not change the expression of ActR-IA, BMPR-IA, BMPR-IB, BMPR-II, and ALK-7 mRNAs. Based on the current data, the CDMPs appear to be able to stimulate the C2C12 cells to differentiate into the osteoblast pathway.

Animals↗
PubMed2005

Cardiac transplantation in patients with iron overload cardiomyopathy.

A review of the published world experience with heart transplantation for iron overload cardiomyopathy (IOC) between 1967 and 2003 as well as review of unpublished cases from the database of the United Network for Organ Sharing since 1992, identified a total of 16 patients (14 men and 2 women). Mean age was 31 years (range, 14-63 years). IOC etiology was hemochromatosis in 11 patients (69%), thalassemia major in 4 (25%), and Diamond-Blackfan anemia in 1 (6%). The 30-day mortality was 12%. Three patients (19%) died within 1 year of the transplant, all of infectious complications. An additional patient died at 7.14 years (unknown cause). The actuarial Kaplan-Meier 1-, 3-, and 5-year survival rates were 81% for all 3 time intervals. The actuarial 10-year survival was 41%.

Adolescent↗
PubMed2004

Identification of an osteogenic protein-1 responsive element in the aggrecan promoter.

Previous studies have demonstrated that osteogenic protein-1 (OP-1), a member of the bone morphogenetic protein (BMP) subfamily of the TGF-beta superfamily, stimulates aggrecan synthesis. To investigate transcriptional regulation of the aggrecan gene by OP-1, we constructed a clone containing a 1 kb region of the 5'-upstream sequence of the mouse aggrecan gene fused to the promoter-less luciferase reporter gene in pGL2-Basic vector. A series of promoter deletion constructs were also generated. Studies of the promoter activity of these DNA constructs in transient-transfected cells revealed that deletion of a 33 bp region rendered the promoter unresponsive to OP-1, BMP-6, and CDMP-1 without affecting BMP-2 and TGF-beta1 responsiveness. Thus, the expression of the mouse aggrecan gene in response to BMPs appears to be the result of a unique combination of different cis-acting elements.

Aggrecans↗
PubMed2004

Cartilage-derived morphogenetic proteins enhance the osteogenic protein-1-induced osteoblastic cell differentiation of C2C12 cells.

Previous studies have shown that osteogenic protein-1 (OP-1; also known as BMP-7) induces differentiation of the pluripotent mesenchymal cell line C2C12 into osteoblastic cells. OP-1 also alters the steady-state levels of messenger RNA (mRNA) encoding for the cartilage-derived morphogenetic proteins (CDMPs) in C2C12 cells. In the present study, the effects of exogenous CDMPs on bone cell differentiation induced by OP-1 in C2C12 cells were examined. Exogenous CDMP-1, -2, and -3 synergistically and dose-dependently enhanced OP-1 action in stimulating alkaline phosphatase (AP) activity and osteocalcin (OC) mRNA expression. AP staining studies revealed that the combination of OP-1 and CDMP enhanced OP-1 action by stimulating those cells that had responded to OP-1 and not by activating additional cells. The combination did not change the mRNA expression of the BMPs and their receptors. CDMP-1 enhanced the suppression of the OP-1-induced expression of the myogeneic differentiation regulator MyoD. CDMP-1 and OP-1 alone stimulated Smad5 protein expression, but the combination of OP-1 and CDMP-1 stimulated synergistically Smad5 protein expression. Thus, one mechanism of the observed synergy involved enhancement of the induced Smad5 protein expression. At the same protein concentration, CDMP-1 is most potent in enhancing OP-1 activity in inducing osteoblastic cell differentiation of C2C12 cells. CDMP-3 is about 80% as potent as CDMP-1, and CDMP-2 is the least potent (about 50% of CDMP-1).

Alkaline Phosphatase↗
PubMed2003

Effects of osteogenic protein-1 (OP-1, BMP-7) on gene expression in cultured medial collateral ligament cells.

Osteogenic protein-1 (OP-1, also called BMP-7), a member of the BMP family and the TGF-beta superfamily, induces formation of new bone and cartilage, but also regulates a wide array of processes. In the present study, the expression of several characteristic biochemical markers of ligaments, such as Six1, Scleraxis, aggrecan, and type I collagen in primary cultures of adult rat medial collateral ligament (MCL) cells was determined. The effects of OP-1 on cell proliferation and on gene expression were subsequently examined. OP-1 stimulated cell proliferation, alkaline phosphatase (AP) activity, and the steady-state mRNA levels of the transcription factor Runx2/Cbfa1 in a dose- and time-dependent manner. The mRNA levels of type I collagen only increased slightly, but the activity of the cloned collagen promoter increased by 2-fold in transiently transfected MCL cells. OP-1 also stimulated aggrecan mRNA expression. The mRNA levels of Six1 and Scleraxis were not detectably altered by OP-1. In control cultures, the steady-state mRNA levels of ActR-I, BMPR-IA, BMPR-IB, and BMPR-II increased as a function of time in culture. The mRNA levels of BMP-1 and -4 increased significantly after 12 days, but those of BMP-2 and -6 did not change. The GDF-1, -3, -5, -6, and -8 mRNA levels in the control cultures also increased as a function of time. OP-1 treatment stimulated mRNA expression of BMPR-IA and BMPR-II, but had little effect on ActR-I and BMPR-IB mRNA expression. OP-1 lowered the BMP-1, -2, and -6 mRNA levels without changing the BMP-4 mRNA level. OP-1 treatment also reduced the mRNA levels of GDFs detected. In summary, the present study demonstrated that OP-1 stimulated cell proliferation and mRNA expression of several biochemical markers in this ligament cell culture model and established the spatial and temporal appearance of several members of the TGF-beta superfamily.

Animals↗
PubMed2003

Inhibition of phosphatidylinositol 3-kinase and p70S6 kinase blocks osteogenic protein-1 induction of alkaline phosphatase activity in fetal rat calvaria cells.

Published studies reveal that Osteogenic Protein-1 (OP-1) and insulin-like growth factor-I (IGF-I) synergistically stimulate alkaline phosphatase (AP) activity and bone nodule formation in fetal rat calvaria (FRC) cells. In the present study, we examined whether there are interactions between the signal transduction pathways activated by these two growth factors. OP-1 did not significantly affect the levels of IRS-1, IRS-2, the p85alpha subunit of phosphatidylinositol 3-kinase (PI 3-kinase) or the extracellular signal-regulated kinase (ERK)-2, but stimulated ERK-1 protein by twofold. OP-1 also induced phosphorylation of ERK-1 and -2, but not of Akt/protein kinase B (PKB), a protein kinase that is downstream of PI 3-kinase. By comparison, IGF-I increased the levels of the phosphorylated forms of ERK-1 and -2, and Akt/PKB. Inhibition of ERK activation by PD98059 did not significantly alter the stimulation of AP activity by OP-1 or OP-1 in combination with IGF-I. In contrast, inhibition of PI 3-kinase activity by LY294002 blocked the induction of AP activity by OP-1 and OP-1 plus IGF-I. Treatment of cells with rapamycin, an inhibitor of the mammalian target of mTOR, resulted in a 47% and a 53% decrease in the AP activity induced by OP-1 alone and by OP-1 plus IGF-I, respectively. These studies suggest that PI 3-kinase and mTOR contribute to the induction of AP activity by OP-1 and the synergistic effect of OP-1 and IGF-I on AP activity in FRC cells.

Alkaline Phosphatase↗
PubMed2003

N-Phenyl-N-purin-6-yl ureas: the design and synthesis of p38alpha MAP kinase inhibitors.

The design, synthesis and SAR of a series of 2,6,9-trisubstituted purine inhibitors of p38alpha kinase is reported. Synthetic routes were devised to allow for array synthesis in which all three points of diversity could be facilely explored. The binding of this novel series to p38alpha kinase, which was predicted to have several key interactions in common with SB-203580, was confirmed by X-ray crystallography of 19 (p38 IC(50)=82 nM).

Crystallography, X-Ray↗
PubMed2003

p38 MAP kinases: key signalling molecules as therapeutic targets for inflammatory diseases.

The p38 MAP kinases are a family of serine/threonine protein kinases that play important roles in cellular responses to external stress signals. Since their identification about 10 years ago, much has been learned of the activation and regulation of the p38 MAP kinase pathways. Inhibitors of two members of the p38 family have been shown to have anti-inflammatory effects in preclinical disease models, primarily through the inhibition of the expression of inflammatory mediators. Several promising compounds have also progressed to clinical trials. In this review, we provide an overview of the role of p38 MAP kinases in stress-activated pathways and the progress towards clinical development of p38 MAP kinase inhibitors in the treatment of inflammatory diseases.

Animals↗
PubMed2002

Restraint of proinflammatory cytokine biosynthesis by mitogen-activated protein kinase phosphatase-1 in lipopolysaccharide-stimulated macrophages.

Exposure of macrophages to LPS elicits the production of proinflammatory cytokines, such as TNF-alpha, through complex signaling mechanisms. Mitogen-activated protein (MAP) kinases play a critical role in this process. In the present study, we have addressed the role of MAP kinase phosphatase-1 (MKP-1) in regulating proinflammatory cytokine production using RAW264.7 macrophages. Analysis of MAP kinase activity revealed a transient activation of c-Jun N-terminal kinase (JNK) and p38 after LPS stimulation. Interestingly, MKP-1 was induced concurrently with the inactivation of JNK and p38, whereas blocking MKP-1 induction by triptolide prevented this inactivation. Ectopic expression of MKP-1 accelerated JNK and p38 inactivation and substantially inhibited the production of TNF-alpha and IL-6. Induction of MKP-1 by LPS was found to be extracellular signal-regulated kinase dependent and involved enhanced gene expression and increased protein stability. Finally, MKP-1 expression was also induced by glucocorticoids as well as cholera toxin B subunit, an agent capable of preventing autoimmune diseases in animal models. These findings highlight MKP-1 as a critical negative regulator of the macrophage inflammatory response, underscoring its premise as a potential target for developing novel anti-inflammatory drugs.

Animals↗
PubMed2002

Interleukin-1F7B (IL-1H4/IL-1F7) is processed by caspase-1 and mature IL-1F7B binds to the IL-18 receptor but does not induce IFN-gamma production.

We have recently reported the identification of four novel members of the interleukin-1 (IL-1) family which we designated as IL-1 homologue 1-4 (IL-1H1-4). These proteins exhibit significant sequence homology to other members of the IL-1 family. Of these homologues, only IL-1H4 (renamed IL-1F7b) was predicted to contain a propeptide domain and a caspase cleavage site. We now report that caspase-1 cleaves IL-1F7b at the predicted site to generate mature IL-1F7b. Caspase-4 was also able to process IL-1F7b, albeit inefficiently. Other caspases and Granzyme-B did not cleave IL-1F7b. Furthermore, adenovirus-mediated expression of IL-1F7b in HEK 293 cells led to in situ processing and secretion of mature IL-1F7b. In a screen to identify a potential receptor, both pro and mature IL-1F7b bound to the soluble IL-18 receptor alpha-Fc (IL-18Ralpha-Fc) but not to the soluble IL-1R-Fc or ST2R-Fc fusion proteins. Mature IL-1F7b bound to the IL-18Ralpha-Fc protein with higher affinity than the pro form, although the affinities for both proteins were significantly lower than that observed for IL-18. Consistent with this observation, only IL-18 and not IL-1F7b induced IFN-gamma production by KG1a cells. We also report that pro and mature IL-1F7b form homodimers with association constants of 4 microM and 5 nM, respectively, suggesting biological relevance to IL-1F7b processing. Finally, we have localized the expression of IL-1F7b protein in discrete cell populations including plasma cells and tumor cells. These data suggest that IL-1F7b may be involved in immune response, inflammatory diseases and/or cancer.

Amino Acid Sequence↗