Chronic migraine-cephalgia related to trigeminal artery: a case report of innovative regenerative approach.

Persistent trigeminal artery (PTA) originates from the posterior bend or lateral wall of the intra-cavernous carotid artery and is the most common occurring type of remnant primitive fetal arteries. In literature, there is limited number of reports on migraine-cephalgia (MC) associated with coexisting PTA. The primitive anastomose arteries that fully belong to the intracranial arterial vascular system are not supposed to perform any supportive functional activity; usually they are subjected to normal biological decay caused by the aging process and metabolic dysfunctions. The hypothesis suggests that these primitive fetal arteries such as PTA may not undergo a fast and structural deterioration but they might be active contributors to a series of mechanisms that can cause a variety of idiopathic complaints. Consequently this would bring a different therapeutic approach other than their surgical removal, which is the accepted option today as a solution for these problems. In this case report, a chronic unilateral MC due to coexisting PTA adjacent to trigeminal nerve is presented. The caliber and location of the PTA was confirmed by a CT-Angiography. The MC treatment was achieved by administration of bio-identical testosterone, human placenta extract (HPE), b-nicotinamide adenine dinucleotide (NADH) and low dose amlopidine.

Age-related declines in α-Klotho drive progenitor cell mitochondrial dysfunction and impaired muscle regeneration.

While young muscle is capable of restoring the original architecture of damaged myofibers, aged muscle displays a markedly reduced regeneration. We show that expression of the "anti-aging" protein, α-Klotho, is up-regulated within young injured muscle as a result of transient Klotho promoter demethylation. However, epigenetic control of the Klotho promoter is lost with aging. Genetic inhibition of α-Klotho in vivo disrupted muscle progenitor cell (MPC) lineage progression and impaired myofiber regeneration, revealing a critical role for α-Klotho in the regenerative cascade. Genetic silencing of Klotho in young MPCs drove mitochondrial DNA (mtDNA) damage and decreased cellular bioenergetics. Conversely, supplementation with α-Klotho restored mtDNA integrity and bioenergetics of aged MPCs to youthful levels in vitro and enhanced functional regeneration of aged muscle in vivo in a temporally-dependent manner. These studies identify a role for α-Klotho in the regulation of MPC mitochondrial function and implicate α-Klotho declines as a driver of impaired muscle regeneration with age.

Effects of secreted factors in culture medium of annulus fibrosus cells on microvascular endothelial cells: elucidating the possible pathomechanisms of matrix degradation and nerve in-growth in disc degeneration.

OBJECTIVE: To test whether the interaction between annulus fibrosus cells (AFCs) and endothelial cells (ECs) disrupts matrix homeostasis and stimulates production of innervation mediators. METHODS: Human microvascular ECs were cultured in the conditioned media of AF cell culture derived from degenerated human surgical specimen. Matrix-metalloproteinases (MMPs) and platelet-derived growth factor (PDGF) of ECs of this culture were analyzed by qRT-PCR, Western, and immunofluorescence. Vascular endothelial growth factor (VEGF), Interleukin-8 (IL-8), and nerve growth factor (NGF) in the media of this cell culture were assayed by ELISA. To determine the effects of ECs on AFCs, qRT-PCR was performed to determine mRNA levels of collagen I, II and aggrecan in AFCs cultured in EC conditioned media. RESULTS: Compared to ECs cultured in naïve media, ECs exposed to AFC conditioned media expressed higher mRNA and protein levels of key biomarkers of invasive EC phenotype, MMP-2 (2×), MMP-13 (4×), and PDGF-B (1.5-2×), and NGF (24.9 ± 15.2 pg/mL vs 0 in naïve media). Treatment of AF cells with EC culture conditioned media decreased collagen type II expression two fold. Considerable quantities of pro-angiogenic factors IL-8 (396.7 ± 302.0 pg/mL) and VEGF (756.2 ± 375.9 pg/mL) were also detected in the conditioned media of untreated AF cell culture. DISCUSSION: AFCs from degenerated discs secreted factors which stimulated EC production of factors known to induce matrix degradation, angiogenesis, and innervation. IL-8 and VEGF maybe the secreted factors from AFCs which mediate a pro-angiogenic stimulus often implicated in the development of disc degeneration.

p38 MAPK inhibition selectively mitigates inflammatory mediators and VEGF production in AF cells co-cultured with activated macrophage-like THP-1 cells.

OBJECTIVES: Recent data have suggested that macrophages are involved in the pathogenesis of discogenic back pain and enhance the secretion of inflammatory mediators in co-cultured annulus fibrosus (AF) cells. The purpose of these studies is to determine the role of p38 mitogen-activated protein kinase (p38 MAPK) signaling in the interactions between macrophage and AF cells. METHODS: Human AF cells were co-cultured with phorbol myristate acetate-stimulated macrophage-like THP-1 cells with and without p38 MAPK inhibition. Conditioned media from co-cultured cells were assayed for interleukin (IL)-6, IL-8, prostaglandin E2 (PGE2), PGF2alpha, and vascular endothelial growth factor (VEGF). Naïve and macrophage-exposed AF cell responses to 10ng/ml tumor necrosis factor-alpha (TNF-alpha) were compared using the same outcome measures. RESULTS: IL-6, IL-8, PGE2, PGF2alpha, and VEGF were secreted in greater quantities by cells maintained in co-culture compared to macrophages or AF cells cultured alone. SB202190 blunted IL-6, PGE2, and PGF2alpha production in a dose-dependent manner in co-culture. However, it did not suppress IL-8 and VEGF production. TNF-alpha-stimulated AF cell inflammatory mediators were up-regulated by macrophage exposure. SB202190 successfully suppressed IL-6, IL-8, PGE2, and PGF2alpha secretion in macrophage-exposed AF cells in response to TNF-alpha. CONCLUSIONS: Annular injury can result in macrophage infiltration, and this can cause enhanced inflammatory mediator and VEGF production by AF cells. The p38 MAPK pathway signals are responsible for much of IL-6 and PG secretion from AF cells with macrophage-like cells, suggesting that blockade of this signal may serve as a therapeutic approach to discogenic pain.

Escherichia coli transcript cleavage factors GreA and GreB stimulate promoter escape and gene expression in vivo and in vitro.

The process of RNA chain initiation by RNA polymerases plays a central role in the regulation of transcription. In this complex phase of transcription, short oligomers are synthesized and released from the enzyme-promoter complex in a reaction termed abortive initiation. The polymerase undergoes many cycles of abortive initiation prior to completion of the initiation process, which is signaled by the translocation of the enzyme away from the promoter, release of sigma factor, and formation of an elongation complex in which the RNA is stably bound. We have studied the parameters that affect escape from the promoter by Escherichia coli RNA polymerase for the phage T7 A1 promoter, the phage T5 N25 promoter, and the chimeric promoter T5 N25antiDSR. The latter site contains a synthetic initial transcribed region that reduces its ability to synthesize RNA both in vivo and in vitro. Clearance from T5 N25antiDSR can be stimulated up to 10-fold in vitro by addition of the E. coli transcript cleavage factor GreA or GreB, but these factors have little effect on transcription from the normal T7 A1 or T5 N25 promoters. Using an E. coli strain lacking GreA and GreB, we were also able to show stimulation of transcription by the Gre factors from the T5 N25antiDSR promotor in vivo. The stimulation of RNA chain initiation by Gre factors, together with their known biochemical properties in the transcription elongation reaction, suggests some specific models for steps in the transcription initiation reaction.

Designing zinc-finger ADR1 mutants with altered specificity of DNA binding to T in UAS1 sequences.

Yeast ADR1 contains two Cys2,His2 zinc fingers needed for DNA binding to the upstream activation sequence UAS1, with bases T5T6G7-G8A9G10 in the ADH2 promoter. Potential DNA-contacting amino acid residues at -1, +3, and +6 in the alpha-helical domains of ADR1's fingers one and two include RHR-RLR; however, the latter finger two residues Leu146 and Arg149 had not proved to be crucial for ADR1 binding, even though Leu146-T6 and Arg149-T5 interactions with UAS1 DNA were predicted. We altered Leu146 or Arg149 by PCR cassette mutagenesis, to study ADR1 mutant binding to 16 UAS1 variants of thymine bases T5 and T6. Mutation of Leu146 to His, making finger two (RLR) like finger one (RHR), decreased binding to wild type UAS1 having T6, but enhanced its binding strength to sequences having purines G6 or A6, similar to binding seen between finger one's His118 and base A9 of UAS1. Mutating Leu146 to Lys caused this finger two RKR mutant to bind strongly to both G6 and T6, possibly by lysine's amine H-bonding to the carbonyl of guanine or thymine. Specificity of ADR1 for UAS1 with T6 may thus be due to hydrophobic interaction between Leu146 and the T6 methyl group. ADR1 mutants with either His or Lys in the central +3 residue (146) of zinc finger two, which have Arg149 in the +6 alpha-helical position, bind with UAS1 mutant sequences having G5 very strongly, T5 strongly, A5 intermediately, and C5 weakly.(ABSTRACT TRUNCATED AT 250 WORDS)