Experimental integrative muscular movement technique enhances cervical range of motion in patients with chronic neck pain: a pilot study.

INTRODUCTION: Neck pain presents a tremendous physical and financial burden. This study compared the efficacy of the complementary and alternative medical treatments of integrative muscular movement technique (IMMT) and Swedish massage on neck pain in women of occupation age, the largest demographic group with neck pain. METHODS: A total of 38 women were assigned to IMMT (n=28) or Swedish massage (n=10) in a blinded manner. Both groups received eight 30-minute treatments over 4 weeks. Cervical range of motion (ROM) in flexion, extension, sidebending, and rotation was measured before and after treatment. Each patient's pain was assessed by using an analogue pain scale of 0-10. RESULTS: Compared with the Swedish massage group, patients receiving IMMT experienced a significant increase in ROM in cervical flexion (p<0.001), extension (p<0.001), sidebending (p<0.05), and rotation (p<0.001). Absolute change in pain for IMMT was -1.75 units compared with -0.3 units for Swedish massage (p<0.05). CONCLUSION: Patients receiving the IMMT demonstrated significantly improved cervical ROM in every movement measured compared with Swedish massage. Inclusion of the IMMT in a treatment regimen for chronic neck pain may lead to decreased pain and increased cervical ROM. These positive effects of the IMMT intervention may have a role in enhancing functional outcomes in patients with neck pain.

A novel massage therapy technique for management of chronic cervical pain: a case series.

BACKGROUND: Neck pain is a generalized condition resulting from a complex etiology with presentation of a wide variety of symptoms. Neck pain is most often accompanied by decreased range of motion (ROM), muscle and joint stiffness, and limitations in functional capabilities. This condition may result in significant personal and societal burden. PURPOSE: We evaluated the effectiveness of a novel massage therapy intervention by following the treatment regimen and outcomes of two patients experiencing chronic neck pain. PARTICIPANTS: Two patients (46 and 53 years old) experienced chronic (>5 years) neck pain. Both patients reported pain, limited ROM, and muscle and joint stiffness. Additionally, the first patient reported a lack of sleep, and both patients stated their pain interfered with their quality of life and activities of daily living. INTERVENTION: Patients received the Integrative Muscular Movement Technique (IMMT) intervention approximately twice a week for a total of eight treatments, each approximately 20 minutes in duration. RESULTS: Both patients experienced a reduction in pain and an increase in cervical ROM in flexion, extension, rotation, and sidebending. The first patient also reported an increased ability to sleep. Both patients reported an increased ability to perform activities of daily living, including work-related responsibilities. CONCLUSIONS: For the two patients included in this report, therapist observations and patient reports indicate that inclusion of the IMMT treatment in a treatment regimen for chronic neck pain may lead to decreased pain and increased cervical ROM. These positive effects of the IMMT intervention may have a role in enhancing functional outcomes of these patients.

Tumor necrosis factor-α treatment of HepG2 cells mobilizes a cytoplasmic pool of ERp57/1,25D3-MARRS to the nucleus.

ERp57/PDIA3/1,25-MARRS has diverse functions and multiple cellular locations in various cell types. While classically described as an endoplasmic reticulum (ER) resident protein, ERp57 has a nuclear location sequence (NLS) and can enter the nucleus from the cytosol to alter transcription of target genes. Dysregulation and variable expression of ERp57 is associated with a variety of cancers including hepatocellular carcinoma (HCC). We investigated the dynamic mobility of ERp57 in an HCC cell line, HepG2, to better understand the movement and function of the non-ER resident pool of ERp57. Subcellular fractionation indicated ERp57 is highly expressed in the ER with a smaller cytoplasmic pool in HepG2 cells. Utilizing an ERp57 green fluorescent protein fusion construct created with and without a secretory signal sequence, we found that cytoplasmic ERp57 translocated to the nucleus within 15 min after tumor necrosis factor-α (TNF-α) treatment. Protein kinase C activators including 1,25-dihydroxyvitamin D(3) and phorbol myristate acetate did not trigger nuclear translocation of ERp57, indicating translocation is PKC independent. To determine if an interaction between the rel homology binding domain in ERp57 and the nuclear factor-κB subunit, p65, occurred after TNF-α treatment and could account for nuclear movement, co-immunoprecipitation was performed under control and conditions that stabilized labile disulfide bonds. No support for a functional interaction between p65 and ERp57 after TNF-α treatment was found in either case. Immunostaining for both ERp57-GFP and p65 after TNF-α treatment indicated that nuclear translocation of these two proteins occurs independently in HepG2 cells.

Regulation of expression of 1,25D3-MARRS/ERp57/PDIA3 in rat IEC-6 cells by TGF beta and 1,25(OH)2D3.

We examined the transcriptional regulation of expression of the redox-sensitive Membrane-Associated-Rapid Response, Steroid-binding (1,25D(3)-MARRS) protein specific for 1,25(OH)(2)D(3) in a rat small intestinal cell line, IEC-6, that demonstrates rapid responses to 1,25(OH)(2)D(3). 1,25D(3)-MARRS binds and is activated by 1,25(OH)(2)D(3), but is not itself up-regulated by treatment with 1,25(OH)(2)D(3), nor is there a Vitamin D response element (VDRE) in its proximal promoter. We previously reported that transforming growth factor beta (TGFbeta) increased steady state levels of 1,25D(3)-MARRS transcript and protein approximately two-fold [Rohe B, Safford SE, Nemere I, Farach-Carson, MC. Identification and characterization of 1,25D(3)-membrane-associated rapid response, steroid (1,25D(3)-MARRS)-binding protein in rat IEC-6 cells. Steroids 2005;70:458-63]. To determine if this up-regulation could be attributed to the function of a highly conserved consensus smad 3 binding element present in the proximal promoter of the 1,25D(3)-MARRS gene, we created a promoter-reporter [SEAP] construct that was responsive to TGFbeta (200 pM). Deletion or mutation of the smad 3 element greatly reduced the response of the 1,25D(3)-MARRS promoter to TGFbeta. Subsequent studies found that the smad 3 response element is bound by a protein found in the IEC-6 nuclear extract, most likely smad 3. Interestingly, although 1,25(OH)(2)D(3) alone did not increase expression of the 1,25D(3)-MARRS promoter-reporter, co-treatment of transfected IEC-6 cells with 1,25(OH)(2)D(3) and TGFbeta shifted the dose-response curve to a lower effective concentration (100 pM peptide). We conclude that TGFbeta is a transcriptional regulator of 1,25D(3)-MARRS expression via a functional smad 3 element and that cross-talk with non-classical 1,25(OH)(2)D(3)-stimulated pathways occurs. The findings have broad implications for redox-sensitive signaling phenomena including those that regulate phosphate transport in the intestine.

Identification and characterization of 1,25D3-membrane-associated rapid response, steroid (1,25D3-MARRS)-binding protein in rat IEC-6 cells.

We report the presence of a mammalian equivalent of the avian Membrane-Associated Rapid Response, Steroid (1,25D3-MARRS)-binding protein specific for 1,25(OH)2D3 in a rat small intestinal cell line, IEC-6, that demonstrates rapid responses to the steroid hormone. Identification of transcript and protein was achieved using RT-PCR with several specific primer sets, Western blot analysis with two separate antibodies recognizing distinct regions of the protein, ribozyme knockdown and immunohistochemistry. Promoter analysis of the 1000-bp upstream region of the 1,25D3-MARRS gene in several species revealed the presence of a conserved smad-3 element in the 5' proximal promoter region, but no classical vitamin D response element (VDRE). Treatment of IEC-6 cells with transforming growth factor beta1 (TGFbeta1) increased steady-state levels of 1,25D3-MARRS (mRNA and protein) approximately two-fold over a 24-h period. In contrast, treatment with 1,25(OH)2D3 failed to significantly change 1,25D3-MARRS protein or mRNA levels. Localization studies showed rapid nuclear translocation of a pool of 1,25D3-MARRS protein after 1,25(OH)2D3 treatment, suggesting that the protein is subject to membrane-initiated signal pathway activation. Together these data point to complex interactions between the two important 1,25(OH)2D3 sensitive response systems in intestinal cells, 1,25D3-MARRS protein and the well-studied nVDR, that together work to fine tune intestinal Ca2+ absorption in a variety of avian and mammalian species.

Identification and characterization of 1,25D3-membrane-associated rapid response, steroid (1,25D3-MARRS) binding protein.

1,25-Dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) operates through pharmacologically distinct nuclear receptor-mediated and plasma membrane-initiated mechanisms. The nuclear receptor is well described, but the membrane receptor identity remains unproven. A 66 kDa protein from chick intestinal basolateral membranes was isolated previously and identified as a candidate receptor (now termed 1,25D(3)-MARRS). A chicken cDNA library was screened for clones encoding the N-terminal sequence of 1,25D(3)-MARRS. An exact match was found with an insert containing an open coding region for the full-length candidate 1,25D(3)-MARRS protein. Analysis reveals a 5' untranslated region, a precursor translation product with methionine start site, a signal peptide and a translation product of 505 amino acids prior to translation termination site. Prosite analysis predicts potential sites for phosphorylation by casein kinase II cAMP-dependent kinase, protein kinase C, and tyrosine kinase and an N-myristoylation site with high probability of occurrence. Additionally, two conserved domains capable of interacting with Rel homology domains (RHD) are present. Oligonucleotide primers sets designed to amplify unique regions of the sequence produced amplimers of the predicted size from both chicken and rat intestinal cells. Transcription-translation produced a protein that was recognized in Western blot analysis by Ab099, a polyclonal antibody recognizing the N-terminus of the 66 kDa MARRS protein.

A rapid and simple nonradioactive method for in vitro testing of ribozyme activity.

Ribozymes that target specific messenger RNA transcripts are powerful tools in the emerging fields of functional genomics, proteomics, and metabolomics. We have found that successful in vitro testing greatly increases the likelihood of producing ribozymes with good efficacy in living cells. A rapid and simple nonradioactive method for systematic in vitro testing of ribozyme-cleaving activity is reported. Ribozymes are synthesized enzymatically from double-stranded DNA (dsDNA) oligonucleotides without vector cloning. Substrate target DNA template is cloned into a vector flanked with SP6 and T7 promoters at multiple cloning sites that permit colorimetric screening and ampicillin selection, enhancing the efficiency of the cloning procedure. Ribozyme cleavage products are satisfactorily resolved on 2.0% NuSieve 3:1 agarose (FMC Products, Rockland, ME)/formaldehyde gels by electrophoresis. This method avoids the preparation of polyacrylamide gels. Using this procedure, the ribozyme, target substrate RNA, and ribozyme cleavage products are all easily detected by ethidium bromide staining. Resolution and detection are fast and simple, eliminating the need for either polyacrylamide gel analysis or radiolabeling. The use of RNase inhibitors in the assays is also assessed and discussed.