IF Staining of Human Primary Neurons

Primary Neurons are inputs or raw materials for cell based assays. When cells do not work as promised, there are multiple costs including lost time and potentially flawed data. Neuromics strives to provide easy to culture, potent and cost effective cells. Proving these capabilities is an ongoing activity for us. This includes testing these cells using our markers.

I wanted to share new immunofluorescence images. Here is a link to the protocol: staining primary neurons.

hN2 cells stained with our chicken polyclonal antibody to Vimentin, in red. Islands of Hn2 cells form after 4 days in culture forming beautiful flower like structures. Vimentin is a well established marker of early differentiating neuronal lineage cells. Taken with a 10X objective lens. Blue staining is the nuclear DNA.

hN2 cells grown in culture for 4 days and stained with our chicken polyclonal to MAP2, a marker of neurons. Differentiating cells show strong cytoplasmic staining for MAP2 . Blue stain is DAPI and reveals cell nuclei of some non neuronal cells in this culture.

hN2 cells grown in culture for 4 days and stained with our chicken polyclonal to Neurofilament light or low molecular weight chain NF-L, a marker of neurons. Many of the differentiating cells show strong cytoplasmic and clearly fibrillar staining for NF-L. Blue stain is DAPI and reveals cell nuclei of some non neuronal cells in this culture.

Fear Changes Hippocampus Neuropns

Fear in mice catalyzes rapid accumalation of EphrinB2 pyramidal neurons of the CA1 area.

This study suggests that rapid accumulation of EphrinB2 in hippocampal CA1 neurons is involved in the behavioural and cellular modifications induced by contextual fear conditioning. A similar mechanism does not appear to occur in lateral amygdala neurons, in spite of the robust behavioural and cellular modifications induced in such structure by cued fear conditioning: Antonio Trabalzaa, Sandra Colazingaria, Carmelo Sgobiob, Arturo Bevilacqua. Contextual learning increases dendrite complexity and EphrinB2 levels in hippocampal mouse neurons. Behavioural Brain Research. doi:10.1016/j.bbr.2011.11.008.

Diabetic retinopathy blindness-root causes

Diabetic retinopathy is a leading cause of acquired blindness. This publication from our friends at University of Buenos Aires touches on potential root causes: Diego C. Fernandez, Laura A. Pasquini, Damián Dorfman, Hernán J. Aldana Marcos, Ruth E. Rosenstein. Early Distal Axonopathy of the Visual Pathway in Experimental Diabetes. doi:10.1016/j.ajpath.2011.09.018
" In animals that had been diabetic for 6 weeks, a large increase in astrocyte reactivity occurred in the distal (but not the intraorbital) portion, which coincided with significant axon loss. Moreover, profound myelin alterations and altered morphologic features of oligodendrocyte lineage were observed at the distal (but not the proximal) optic nerve portion. The present results suggest that axoglial alterations at the distal portion of the optic nerve could be the first structural change in the diabetic visual pathway."
The authors used our PDGFR Alpha/CD140A Marker to Study the change in Oligodendrocyte Lineage precursor cells. Expression of the protein was increased in these cells with the presence of disorganized and hypertrophic cells. This could disrupt formation of myelin resulting the pathological alteration at the distal portion.

Opioid Induced Itch

Our widely used and frequently published Opioid Receptors Antibodies are used for a spectrum of pain research. Here is an interesting study on the root causes of opioid induced itch.

Xian-Yu Liu, Zhong-Chun Liu, Yan-Gang Sun, Michael Ross1, Seungil Kim, Feng-Fang Tsai, Qi-Fang Li, Joseph Jeffry, Ji-Young Kim, Horace H. Loh, Zhou-Feng Chen. Unidirectional Cross-Activation of GRPR by MOR1D Uncouples Itch and Analgesia Induced by Opioids. Cell, Volume 147, Issue 2, 14 October 2011, Pages 261-262.

Root Causes: Spinal opioid-induced itch, a prevalent side effect of pain management, has been proposed to result from pain inhibition. We now report that the μ-opioid receptor (MOR) isoform MOR1D is essential for morphine-induced scratching (MIS), whereas the isoform MOR1 is required only for morphine-induced analgesia (MIA). MOR1D heterodimerizes with gastrin-releasing peptide receptor (GRPR) in the spinal cord, relaying itch information. We show that morphine triggers internalization of both GRPR and MOR1D, whereas GRP specifically triggers GRPR internalization and morphine-independent scratching. Providing potential insight into opioid-induced itch prevention, we demonstrate that molecular and pharmacologic inhibition of PLCβ3 and IP3R3, downstream effectors of GRPR, specifically block MIS but not MIA. In addition, blocking MOR1D-GRPR association attenuates MIS but not MIA. Together, these data suggest that opioid-induced itch is an active process concomitant with but independent of opioid analgesia, occurring via the unidirectional cross-activation of GRPR signaling by MOR1D heterodimerization.

Mouse epiSCs Into Myelinating Cells

This study published recently in Nature Methods hit my radar scope becaused it referenced use of our widely used and frequently published stem cell marker Tuj 1 (Neuron-specific class III beta-tubulin): Fadi J Najm, Anita Zaremba, Andrew V Caprariello, Shreya Nayak, Eric C Freundt, Peter C Scacheri, Robert H Miller & Paul J Tesar. Rapid and robust generation of functional oligodendrocyte progenitor cells from epiblast stem cells. Nature Methods (2011) doi:10.1038/nmeth.1712.

Dr. Paul Tesar and his team at Case Western University demonstrated the ability to convert pluripotent epiblast stem cells into pure populations of myelinating cells, called oligodendrocyte progenitor cells (OPCs). First, stem cells in a petri dish are treated with molecules to direct them to become the most primitive cells in the nervous system. To produce OPCs, these primitive cells are treated with a defined set of proteins. The cells were cultured on laminin and treated withh apporopriate growth factors. The OPCs were nearly homogenous and could be multiplied to obtain more than a trillion cells.

The OPCs were treated with thyroid hormone, which is key to regulating the transition of the OPCs to oligodendrocytes. The result was the OPCs stopped proliferating and turned into oligodendrocytes within four days.

These methods could used to potentially produce stable and pure populations of human OPCs in a significant enough number to treat patients with demyelinating diseases such as multiple sclerosis and cerebral palsy.

Immunostaining Neurons and Glia

I would like to thank Dr. Gerry Shaw, University of Florida for his excellent work with our Primary Neurons and Astrocytes and Neuronal-Glial Markers. Here's an example image with many more to follow:

Image: E18 hippocampal neurons stained with MAPT (red) and Doublecortin (green). The two proteins overlap in the proximal dendrites, but doublecortin is more abundant in the growth cones and periphery. As a result, the periphery appears green while the more proximal regions of the cells are yellow. The single longer process of this cell, presumably an axon, has a low doublecortin content and so appears red. Blue staining is the nuclear DNA. Protocol on datasheet.

BDNF and Exercise Study-Running Mice

Get fit and get smart. There is increasing evidence that vigorous exercise increases secretion of Brain-Derived Neurotrophic Factor (BDNF) Protein.  BDNF is a catalyst of processes that increase growth of neurons especially in the hippocampus.

In this study researchers show new cell proliferation, survival, neuron number, and neurotrophin levels were enhanced only when running was accessible to mice. They conclude that exercise is the critical factor mediating increased BDNF levels and adult hippocampal neurogenesis: Tali Kobilo, Qing-Rong Liu, Kriti Gandhi, Mohammed Mughal, Yavin Shaham and Henriette van Praag. Running is the neurogenic and neurotrophic stimulus in environmental enrichment. doi: 10.1101/lm.2283011. Learn. Mem. 2011. 18: 605-609... human recombinant BDNF (0.1 µg) monomer (Neuromics)...
BDNF, CF Recombinant Protein
Related Reagents:
Neuron-Glial Expressed-Includes Neurotrophin Proteins
Neurotrophins and Growth Factor Antibodies
Primary Neurons and Astrocytes-Primary human, rat and mouse neurons and astrocytes.

Keep your brain healthy.

Immune-Inflammatory Response and Pain Research

Our Pain and Inflammation Related Research Antibodies are increasingly being used to study the root causes of immune/inflammatory related pain induction. Here're related publications: Lintao Qu, Pu Zhang, Robert H. LaMotte, Chao Ma. Neuronal Fc-gamma receptor I mediated excitatory effects of IgG immune complex on rat dorsal root ganglion neurons. Brain, Behavior, and Immunity. Volume 25, Issue 7, October 2011, Pages 1399-1407......rabbit-anti-TRPV1, 1:1000, Neuromics...

Highlights: Pain often accompanies antigen-specific immune-related disorders though little is known of the underlying neural mechanisms. A common feature among these disorders is the elevated level of antigen-specific immunoglobulin (Ig) G in the serum and the presence of IgG immune complex (IC) in the affected tissue. We hypothesize that IC may directly activate the Fc-gamma receptor type I (FcγRI) expressed in nociceptive dorsal root ganglion (DRG) neurons and increase neuronal excitability thus potentially contributing to pain. Immunofluorescent labeling indicated that FcγRI, but not FcγRIIB or FcγRIII, was expressed in a subpopulation of rat DRG neurons including those expressing nociceptive markers. Calcium imaging revealed that the IC, but neither of the antibody (IgG) or antigen alone, produced an increase in intracellular calcium. This effect was abolished by the removal of the IgG Fc portion in the IC or the application of an anti-FcγRI antibody, suggesting a key role of the FcγRI receptor. Removal of extracellular calcium or depletion of intracellular calcium stores prevented the IC-induced calcium response. In whole-cell current-clamp recordings, IC depolarized the resting membrane potential, decreased the rheobase, and increased the number of action potentials evoked by a depolarizing current at 2× rheobase. In about half of the responsive neurons, IC evoked action potential discharges. These results suggest that a subpopulation of nociceptive neurons expresses functional FcγRI and that the activation of this receptor by IC increases neuronal excitability.

B. Huanga, X. Zhaoc, L.-B. Zhengb, L. Zhanga, B. Nia. Different expression of tissue inhibitor of metalloproteinase family members in rat dorsal root ganglia and their changes after peripheral nerve injury. Neuroscience, Volume 193, 13 October 2011, Pages 421-428....anti-P2X3 (rabbit, Neuromics, MN, USA)...

Xona Microfluidics and Neurons

I am impressed with these Video from the Jeon Lab at UC Irvine. It represents a novel method for neuro-drug discovery:
Preparing E18 Cortical Rat Neurons for Compartmentalization in a Microfluidic Device.

This technology represents a way to separate axon from cell bodies.

hNP1™ Human Neural Progenitors Video Protocols

Video Protocols designed to insure your success with our hNP1^TM Human Neural Progenitors

hNP1™ Human Neural Progenitors Video Protocols

Thawing, Culturing and Transfer-Sub Culturing Protocols.

Opioid Receptor Antibodies Trifecta

This publication proposes a role for opioid receptors in treating cancers. It also references use of our μ, δ, and κ opioid receptor antibodies.

Kohei Yamamizu1, Sadayoshi Furuta, Shiori Katayama, Michiko Narita, Naoko Kuzumaki, Satoshi Imai, Hiroshi Nagase, Tsutomu Suzuki, Minoru Narita, and Jun K. Yamashita. The κ opioid system regulates endothelial cell differentiation and pathfinding in vascular development. Blood July 21, 2011 vol. 118 no. 3 775-785.

Highlights: The opioid system is, thus, a new regulator of vascular development that simultaneously modifies 2 distinct vascular properties, EC differentiation and vascular pathfinding. We confirmed that KOR, but not MOR, was highly expressed in various ECs such as HUVECs (data not shown), suggesting that KOR agonists could directly act on tumor ECs to suppress VEGF receptor expression, similar to the effects observed in embryonic ECs. If so, a combination therapy including an MOR agonist, morphine, and a KOR agonist (such as TRK820, a clinically approved drug in Japan for uremic pruritus) may prove useful for cancer therapy through the suppression of tumor angiogenesis by dual inhibition of VEGF ligands and receptors, extending the therapeutic benefits beyond pain relief.

Images: KOR was highly expressed in Flk1⁺ vascular progenitors. (A) RT-PCR showing mRNA expression of MOR, DOR, and KOR in ES cells, Flk1⁺ cells, cells after 1 or 3 days of Flk1⁺ cell culture (Flk-d1 or Flk-d3), CD31-positive cells (ECs) and CD31-negative cells (MCs) at Flk-d3. (B) Fluorescent staining for MOR, DOR, and KOR at Flk-d1. Nuclei are stained with DAPI (blue). Left, MOR; middle, DOR; right, KOR. Scale bars, 100 μm. (C) Double fluorescent staining for MOR, DOR, and KOR with CD31 (red) at Flk-d3. Nuclei are stained with DAPI (blue). Top, opioid (green) receptors (green); middle, CD31 (red); bottom, merged. Scale bars, 100 μm.

Is Neuropathy Really Gliopathy?

I found this excellent website from posting by Dr. Jan M. Keppel Hesselink, Professor molecular pharmacology, director Institute neuropathic pain: http://www.neuropathie.nu/. It represents a new way of understanding root causes and potential therapies for Neuropathic Pain. Here're highlights:

Gliopathic pain: is a brand new term for what we always thought to be neuropathic pain. It refers to pain related to neuropathic pain, however, the primary driver of this pain is most probably more linked to glia and asterocytes. The mechanism of gliopathic pain is the hyperactivation of glia cells, which results in neuropathic pain.

The role of Glia and Astrocytes:Glia and astrocytes play a central role in neuropathic pain, and gliopathic pain, or asteropathic pain will become new synonyms for neuropathic pain. In a recent hallmark paper the term 'Gliopathic pain' was coined.

This is a reason to put our magnifying glass on glia. Gliamodulating drugs will become a new class of neuropathic drugs, the so called gliopathic modulating drugs, and the first prototype, the endogenous fatty acid palmitoylethanolamide, has already been explored in positive proof of principle studies.

For more than a century doctors are aware of the special properties of glia in response to injury. In Germany in 1894 professor Franz Nissl decribed the reaction of glial cells in relation to the nerve fibers in the spinal cord and highlighted their morphological changes after injury. Microglia becomes mores bigger and more abundant after injury and these glial responses can be seen as a biological reponse to promote nerve repair after injury. However, this response can go berzerk and might be one of the most important mechanisms leading to neuropathic pain.

This is a short synopsis. There is a wealth of more information on the website. That said, I will be posting more on Gliopathic Pain.

Primary Neurons and Cell Based Assays

The feedback I receive from Neuroscientists is consistent. To paraphrase, "gives us healthy, consistent and potent primary cells. I understand the hard work it takes to generate meaningful and publishable results from cell based assays. Our Primary Neurons and Astrocytes are merely inputs for these assays. The real cost is the time invested in culturing and time lost if they don't work.

I have numerous postings on success: Primary Neurons Postings. I wanted to share more data and feedback.

Primary DRGs-Culturing these can be tricky. I make it a point to work with labs to make sure the protocol options best match the desired outcome for assays. This includes replacing cells to make sure we can accurately troubleshoot. This approach insures I can pin point the issues and make sure they are all resolved in round two. Here's a representative testimonial: "Thanks for following up, the DRGs worked great and we were able to get excellent data from them. Thanks so much for working with us." Adam Ross, Dr. Chengji Zhou Lab, UC Davis

Image: DRGs cultured on Calf Skin Collagen.

Primary Hippocampal Neurons-I would like to thank Vimal Swarup, University of Utah for this excellent image.

The cells have been fixed after 48 hrs, they were grown over poly-lysine coated coverslips in the media supplied by Neuromics. Cells were imaged in phase contrast mode with 40x objective.

Put our primary cells to the test!

MOR and NMDAR Interplay-Implications in Pain Control

Our Opioid Receptor Antibodies continue to be referenced in publications by Pain Researchers. Many of these studies provide a greater understanding of how opioids alleviate pain and what modulates this ability.

For example, the capacity of opioids to alleviate inflammatory pain is negatively regulated by the glutamate-binding N-methyl-D-aspartate receptor (NMDAR). This study drills down into the specifics of this regulation and references use of Neuromics' MOR1C Antibody: María Rodríguez-Muñoz, Pilar Sánchez-Blázquez, Ana Vicente-Sánchez, Esther Berrocoso and Javier Garzón. María Rodríguez-Muñoz, Pilar Sánchez-Blázquez, Ana Vicente-Sánchez, Esther Berrocoso and Javier Garzón. The Mu-Opioid Receptor and the NMDA Receptor Associate in PAG Neurons: Implications in Pain Control. Neuropsychopharmacology , (3 August 2011) | doi:10.1038/npp.2011.155.

Abstract: The capacity of opioids to alleviate inflammatory pain is negatively regulated by the glutamate-binding N-methyl-D-aspartate receptor (NMDAR). Increased activity of this receptor complicates the clinical use of opioids to treat persistent neuropathic pain. Immunohistochemical and ultrastructural studies have demonstrated the coexistence of both receptors within single neurons of the CNS, including those in the mesencephalic periaqueductal gray (PAG), a region that is implicated in the opioid control of nociception. We now report that mu-opioid receptors (MOR) and NMDAR NR1 subunits associate in the postsynaptic structures of PAG neurons. Morphine disrupts this complex by protein kinase-C (PKC)-mediated phosphorylation of the NR1 C1 segment and potentiates the NMDAR–CaMKII, pathway that is implicated in morphine tolerance. Inhibition of PKC, but not PKA or GRK2, restored the MOR–NR1 association and rescued the analgesic effect of morphine as well. The administration of N-methyl-D-aspartic acid separated the MOR–NR1 complex, increased MOR Ser phosphorylation, reduced the association of the MOR with G-proteins, and diminished the antinociceptive capacity of morphine. Inhibition of PKA, but not PKC, CaMKII, or GRK2, blocked these effects and preserved morphine antinociception. Thus, the opposing activities of the MOR and NMDAR in pain control affect their relation within neurons of structures such as the PAG. This finding could be exploited in developing bifunctional drugs that would act exclusively on those NMDARs associated with MORs.

I will continue to post these studies. They give hope for pain sufferers as many propose potential new druggable targets.

Plasma netrin-1 is a diagnostic biomarker of human cancers

I am pleased to report broadening application for our Stem Cell Markers as a diagnostic for cancers. This publication references use of our Netrin-1 antibody.

Ganesan Ramesh, Arthur Berg, and Calpurnia Jayakumar. Plasma netrin-1 is a diagnostic biomarker of human cancers. Biomarkers. Author manuscript; available in PMC 2011 July 26. Published in final edited form as: Biomarkers. 2011 March; 16(2): 172–180. Published online 2011 February 8. doi: 10.3109/1354750X.2010.541564.
Objectives: To determine whether plasma netrin-1 can be used as a diagnostic biomarker of human cancer.

Materials and Methods: A total of 300 cancer plasma samples from breast, renal, prostate, liver, meningioma, pituitary adenoma, glioblastoma, lung, pancreatic and colon cancer patients were compared against 138 control plasma samples. Netrin-1 levels were quantified by ELISA and immunohistochemistry.

Results: Plasma netrin-1 levels were significantly increased in breast, renal, prostate, liver, meningioma, pituitary adenoma, and glioblastoma cancers as compared to control samples.

Discussion and Conclusion: Our results suggest that plasma netrin-1 can be used as a diagnostic biomarker for many human cancers.

Image: Immunohistochemical localization of netrin-1 in renal cell carcinoma (RCC) tissues. A. Secondary antibody control showing no staining. B. Normal adjacent tissues do not show any staining for netrin-1. C. Stage I RCC shows staining for netrin-1. D. Stage II RCC shows staining for netrin-1. E-F.


Netrin-1 Immunohistochemistry: Stage I–III renal cell carcinoma and normal tissue section (Tissue Array) was obtained from Biomax to immunolocalize netrin-1, as described previously (29). Briefly, tissue sections were dewaxed and rehydrated with graded ethanol (100%, 90%, 70% and 30%) and then washed with PBS. Antigen retrieval was carried out using citrate buffer and steamer. The tissue section was permeabilized with 0.2% Triton X-100 in PBS, and washed and blocked with PBS containing 5% donkey serum and 1% BSA. Primary antibodies included a chicken anti-netrin-1 polyclonal antibody (Neuromics cat # CH23002). Primary antibodies were detected using secondary antibodies conjugated with biotin, which was followed by incubation with streptavidin-horseradish peroxidase (Pierce). Slides were mounted in permount and photographed using an Olympus microscope attached to a CCD camera.
Potent diagnostic tools for cancers save lives. This is especially true of markers that can diagnose them in early stages. I will post these important studies as the cross my radar scope.

SOX2 and Initiation of Breast Tumors

I consider it a feather in Neuromics' cap when our reagent(s) are referenced in a Nature Journal. More importantly it enables me to keep the pulse on novel and important discovery.

This pub references use of one of our SOX2 Antibody and comes from Dr. Angel García Martín and his Team at INBIOMED: O Leis, A Eguiara, E Lopez-Arribillaga, M J Alberdi, S Hernandez-Garcia, K Elorriaga, A Pandiella, R Rezola and A G Martin. Sox2 expression in breast tumours and activation in breast cancer stem cells. Oncogene , (8 August 2011) | doi:10.1038/onc.2011.338.

The important insight from this study is: "Over-expression of Sox2 increased mammosphere formation, effect dependent on continuous Sox2 expression; furthermore, Sox2 knockdown prevented mammosphere formation and delayed tumour formation in xenograft tumour initiation models. Induction of Sox2 expression was achieved through activation of the distal enhancer of Sox2 promoter upon sphere formation, the same element that controls Sox2 transcription in pluripotent stem cells. These findings suggest that reactivation of Sox2 represents an early step in breast tumour initiation, explaining tumour heterogeneity by placing the tumour-initiating event in any cell along the axis of mammary differentiation."

Could these findings ultimately lead to a better diagnostic for Breast Cancer? I'll keep you posted.

Lab Highlights: Breast cancer is the most frequent cancer in women making up to 20% of all tumours diagnosed in women, with 1 million new cases diagnosed every year worldwide (16,000 new cases only in Spain). Worlwide it causes over 350,000 deaths with an increasing tendency. Breast cancer stem cells show the phenotype CD44+/CD24low/-Lin- though only a fraction of this population has the capacity to initiate tumours. Therefore a complete and precise description of the breast cancer stem cells is lacking.

The focus of this laboratory is to identify, isolate and culture breast cancer stem cells from natural breast tumours and compare at the molecular level with normal mammary stem cells. This research involves the stablishment of both in vitro and in vivo functional assays and the molecular characterization (both genomic and proteomic) of breast cancer stem cells to define the mechanisms responsible for its transformed phenotype.

High Content and High Throughput Toxicity Screening

Kits designed for Drug Discovery and Development

Our customers have been impressed with the capablities of our in vivo and in vitro apoptosis and toxicity kits. Here's a recent pub referencing use of one of our FLICA™ in vitro Caspase Kits: Giovanna Grandinetti, Nilesh P. Ingle, and Theresa M. Reineke. Interaction of Poly(ethylenimine)–DNA Polyplexes with Mitochondria: Implications for a Mechanism of Cytotoxicity. Mol. Pharmaceutics, Article ASAP Publication Date (Web): June 23, 2011 Copyright © 2011 American Chemical Society...inhibitor of caspases (FLICA) specific for caspase-9 (Neuromics, Inc)...
Images: Jurkat cells were treated with 1 µM staurosporine for 3 hours to induce caspase 9 activity (top), or were treated with a control (bottom). Both populations were incubated with ICT’s green FAM-LEHD-FMK FLICA™ caspase 9 reagent. DIC images were taken of both samples. Almost all cells in the induced sample (top) fluoresce green therefore they have activated caspase 9. None of the control cells (bottom) fluoresce green, therefore they do not have activated caspase 9 Courtesy of Dr. Brian Lee, ICT

Neuromics is pleased to announce the addition of HemoGenix® Predictive in vitro Toxicity and Apoptosis Kits:

Solutions for predictive in vitro toxicity and apoptosis are now important than ever. These kits enable you to do high throughput and high content screening of stem cells, progenitors and primary cells.
Kit options include:

•LumiSTEM™-96 iPS and LumiSTEM™-iPS HT Assays to Study Induced Pluripotent Stem Cells (iPS) and Toxicity to iPS Cells and Cells Derived from iPS Cells.

•LUMENESC™-Tox HT (LUMENESC™-96 Tox and LUMENESC™-384 HT). A Toxicity Screening and Testing Platform for Cells of the Mesenchymal Stem/Stromal Cell System.

•HALO®-Tox HT Predictive Hemotoxicity Platform using HALO®-96 Tox and HALO®-384 HT. A Highly Predictive, In Vitro Stem and Progenitor Cell Hemotoxicity Screening and Testing Platform for all Stages of Drug Development and Xenobiotic.

I will continue to post regarding progress.

TRPV1 and Diabetic Neuropathy

Thermal hyperalgesia is a common sympton of Diabetic Periperal Neuropathy (DPN). It is one of most difficult types of pain to treat. The development of tolerance, inadequate relief and potential toxicity of classical antinociceptives warrant the investigation of the newer agents to relieve this pain.

The elevated expression of Transient receptor potential vanilloid 1 (TRPV1) suspected as a transmitter of this pain. Dr. Louis Premkumar and his team at SIU have recently published results that further demonstrate the role of TRPV1: Mahendra Bishnoi, Christine A Bosgraaf, Mruvil Abooj, Linlin Zhong, Louis S Premkumar. Streptozotocin-Induced Early Thermal Hyperalgesia is independent of Glycemic State of Rats: Role of Transient Receptor Potential Vanilloid 1(TRPV1) and inflammatory mediators. Molecular Pain 2011, 7:52 doi:10.1186/1744-8069-7-52. Published: 27 July 2011.

Figure 4. Altered TRPV1 staining in spinal cord dorsal horn of STZ-treated rats. A. Representative images of TRPV1 staining from a vehicle-treated, STZ-HG and STZNG rats. An enlarged segment has also been shown. B. Average gray values/10,000 μm² area of TRPV1 staining in dorsal horn was significantly increased (p<0.05) in both STZ-HG and STZ-NG rats as compared to vehicle-treated rats. Asterisk (*) represents p < 0.05. Scale bar is 200 μm and 50 μm for upper and lower panels, respectively.

Conclusions: From these results, it is concluded that TRPV1 is an integral component of initiating and maintaining inflammatory thermal hyperalgesia, which can be alleviated by intrathecal administration of RTX. Further, the results suggest that enhanced expression and inflammation-induced sensitization of TRPV1 at the spinal cord may play a role in central sensitization in STZ-induced neuropathy.

Therapies that downregulate or silence TRPV1 expression could be the key to better treatments for the Thermal Algesia cause by diabetes. I will keep you posted.

hNP1 Neural Progenitors Thawing+ Plating Protocol

This is a great video that shows thawing + plating protocol for our eSC Derived Human NP1 Neural Progenitors.

Potent and Cost Effective Cell Based Assays

I have had many conversations with basic and drug discovery researchers on improving cell based assays. Here's the wish list:

• More potent cells/media
• More accurate analytic tools-quatititative and reproducible results
• Ability to use cells and tools in high throughput/high content screening.
• Cost effectiveness

This wish list is front and center in determining the cells/media and related tools we add to Neuromics' offerings. We are pleased to announce the addition of our Hemogenix's Bioluminomics™ In-Vitro Cell Assays, MSCGro™ Mesenchymal Stem Cell Media and Umbilical Cord Blood derived hMesenchymal Stem Cells.

These provide quantitation, not subjectivity. It includes assay calibration and standardization. It means assay validation. It produces results you can trust and rely on. It means innovation and flexibility. It is advanced technology that is fast to learn, easy to use and above all, cost effective.

Assays options:

• LumiSTEM™ for Primary Stem and Explanted Cells, Stem and Other Cell Lines-Selecting LumiSTEM™ Assays(pdf - 418Kb)
• LUMENESC™ for Mesenchymal Stem/Stromal Cell System-Selecting LUMENESC™ Assays(pdf - 984Kb)
• HALO® for for Human Lympho-Hematopoietic Stem and Progenitor Cell Research.-Selecting HALO™ Assays(pdf - 1.7M)
• Biolumonics Basic Procedures(pdf - 323Kb) 

Available Cells:

Primary Neurons and Astrocytes-Primary human, rat and mouse neurons and astrocytes
STEMEZ^TM Human Neural Progenitor Neuron Discovery Kits-Derived from H9 (WA09) ECSs-Consistent, Easy to Use & Cost Effective
Human Mesenchymal Stem Cells (hMSCs-hMSCs derived from pancreas and umbilical cord blood
Mammalian Cell Lines

Media:
STEMEZ(TM) hN2 Human Neurons Culture Media
MSCGro™ Mesenchymal Stem Cell Media
NbActiv4

I will continue to post customer input and related data on Neuromics' Cell Based Assay Tools.