T cells are involved with ischemia reperfusion injury to the kidney

This article examined the role of T cells in acute renal failure (ARF). In vitro, ARF can be studied by performing bilateral ischemia (BI) on both kidneys in mice. This is done by clamping both the right and left kidney peduncles for around 22 minutes, restricting blood flow to both kidneys. After 22 minutes, the clamps are removed, and blood flow returns to the kidney.

Of course, ischemia-reperfusion injury results from doing BI on the kidneys. This causes pro inflammatory cytokines to be released throughout the blood, thus damaging other parts of the body. Recently, it was found that T cells are directly involved with ischemia reperfusion injury (Savransky et. Al, 2006).

Researchers in this article tested their hypothesis by looking at mice with deficient T cell receptors (TCR). They found that mice lacking the TCR were essentially protected against ischemia reperfusion injury; specifically, deficient TCR mice had a lower level of TNF-α and IL-6 (Savransky et. Al, 2006). The researchers used techniques like flow cytometry to confirm their TCR deficient mice were truly deficient, as well as an myeloperoxidase (MPO) assay to determine if neutrophils and macrophages had infiltrated the kidney tissue. However, they found no significant difference in neutrophil and macrophage infiltration at 24 hr between WT and TCR deficient mice (Savransky et. Al, 2006).

I personally found this article interesting because of the nature of my work. I do research on kidney failure and I thought it was great to find an article that correlates between my research and the topics of this class. I’m curious to know if anyone has further tested the data these researchers found by depleting macrophages in WT mice. Specifically, if you inject WT mice with LEC (liposome encapsulating clodronate, a macrophage ‘killer’) and then perform BI in the same fashion as these researchers, would you also get decreased TNF-α and IL-6 results?

Reference

“Role of the T-cell receptor in kidney ischemia-reperfusion injury.”

V Savransky, R R Molls, M Burne-Taney, C-C Chien, L Racusen and H Rabb.

Kidney International (2006) 69, 233-238.

Toxicity of NSAID's in Rat Models

As we read a couple weeks ago in the review articles about the ‘Use of Non Steroidal Anti-inflammatory Drugs’ NSAID’s are very controversial in clinical use as they have shown increase use risk of cardiovascular and cerebrovascular events. This article was very good review article to warn people of the clinical cases that have been reported but there was no lab research done on these drugs. The article “Comparison of the Intestinal Toxicity of Celecoxib, a Selective COX-2 Inhibitor, and Indomethacin in the Experimental Rat” gives a new perspective on NSAID’s and compares two NSAID’s, Celecoxib a selective COX-2 inhibitor and Indomethacin a non-selective COX inhibitor. The claim is that all the problems with NSAID’s are that they cause a ‘topical’ effect and inhibit mucosal constitutive COX-1 enzyme. They used both in vivo and in vitro methods of Celecoxib and Indomethacin to test intestinal permeability and inflammation along with pro inflammatory proteins and production of ulcers. They found that Indomethacin, a non-selective COX inhibitor had a significant damaging effect to the intestinal mucosa, where Celecoxib, a selective COX inhibitor showed less intestinal damage, lower PGF levels and did not show the ‘topical’ effect on the mitochondria.
This article in essence showed a in vivo and in vitro with rat models and found evidence that shows that different NSAID’s may have different effects. Granted that this study was more focused on the digestive system and most of the problems reported have been with the cardiovascular and cerebrovascular systems. It does shed a new light on NSAID’s and maybe the idea to look at research of NSAID’s effect on the heart and brain.

Comparison of the Intestinal Toxicity of Celecoxib, a Selective COX-2 Inhibitor, and Indomethacin in the Experimental Rat
J.A. Tibble, G. Sigthorsson, R. Foster, I Bjarnason

Turmeric Supplements Shown To Prevent Rheumatoid Arthritis Flares

As I last blogged, Turmeric has been used for centuries for treatment of inflammatory disorders. It seems as if it is a too good to be true cure all for numerous ailments though, as there is not much information regarding its true efficiency. The research article entitled "Turmeric Extracts Containing Curcumoids Prevent Experimental Rheumatoid Arthritis" sheds new light on turmeric stating that it does have an antiarthritic effect in Rheumatoid Arthritis (RA). However, do not bother reading the article for yourself because it is actually quite difficult to decipher from which turmeric extract they are speaking of through out the article's entirety. So I will just give you the condensed version!
The curcuminoids extracted from turmeric are believed to inhibit the production of the cytokines TNF-alpha, and IL-1B along with PGE2. They are thought to be inhibitors of transcription factor activation such as the activation of NF-kB. If this is activated during a bout of inflammation it will inevitably lead to increased expression of cytokines, chemokines, and COX-2 (inflammatory protein), which are the mediators of joint inflammation in RA. Random over the counter supplements of turmeric were chosen and then tested to find that the majority of them were actually composed of less than 50% curcuminoids. The researchers prepared and chemically characterized a complex turmeric extract depleted in essential oils and composed of less than 50% curcuminoids, isolating it from a commercial source of turmeric rhizome, and then tested it for the in vitro antiinflammatory efficiency along with its in vivo antiarthritic efficiency. The efficiency of this oil-depleted turmeric fraction was then compared with commercial curcumin product (which have greater than 90% curcuminoids). The animal model used in the experiment was a streptococcal cell wall (SCW) induced arthritis, which is the animal model for rheumatoid arthritis. Over a 28 day time course, female Lewis rats having SCW developed an acute phase of joint swelling to be followed by a chronic phase of inflammation that was characteristic of actual joint destruction.
The oil-depleted turmeric extract (40.6% curcuminoid by weight) was administered intraperitoneally daily starting 4 days before the SCW administration. This was shown to significantly inhibit joint inflammation in the acute inflammatory phase by 64% and the chronic destructive phase of arthritis was inhibited 72%. Alternatively, if treatment was administered after the acute phase but before the start of the chronic phase, the same dose did not have any effect on joint swelling. Also, as would be expected, four days before administering SCW treatment with a lower dose of the turmeric fraction showed no effect on joint swelling.
In contrast to how ineffective a low dose of turmeric fraction was on joint swelling, administration with the same lose does of purified curcuminoids (93.6% curcuminoid by weight) starting four days prior to SCW administration significantly inhibited joint inflammation in both acute and chronic phases of arthritis (75% and 68% inhibition). The turmeric fraction dose had to be increased by six-fold in order to have these same effects. The remaining components of the turmeric fraction , besides the three major curcuminoids, may actually be preventing the antiarthritic effect of the curcuminoids.
A granulomatous inflammatory response occurs in the liver and spleen of the rats at the sites of SCW deposition. A high does of turmeric fraction prevented hepatic granulomas. A low dose of purified curcuminoids had no effect on the formation of granuloma formation. The fact that a high dose of the turmeric fraction inhibited granuloma formation most like indicates that a higher dosage of curcuminoids is necessary to induce an antiinflammatory effect.
Essentially this study provides evidence that turmeric may actually be a legitimate antiinflammatory agent. Turmeric products seem to be useful only for the prevention of arthritis but not for treatment during active inflammation. I will be interested to see in the next couple of years what further research states about turmeric. But what do you think...do you guys believe that this is enough evidence for further clinical testing of curcuminoids from turmeric supplements for inhibiting RA flares?



Turmeric Extracts Containing Curcuminoids Prevent Experimental Rheumatoid Arthritis
Janet L. Funk, Janice N. Oyarzo, Jennifer B. Frye, et al.

Ebola Virus Can Recombine

Ebola virus was discovered in 1976 and has since, swept across Africa causing fever, hemorrhage and death in up to 90% of people who are infected. Just this last August, 76 people were affected in the Democratic Republic of Congo. This disease is equally fatal to ape populations. The lowland gorilla is now classified as an endangered species due to multiple outbreaks over the last few years.

Research comparing recent samples from six gorillas and a chimpanzee revealed that the virus from all animals have similar genes. The genes are also homologous to Ebola virus obtained from human samples in the region. However, when compared to a broader range of Ebola virus samples, the virus sequences map to two separate strains. Gene sequences of Ebola virus in human samples taken before 1996 are all more similar to each other than to viral sequences from human and ape samples since then.

This in itself suggests mutation of the virus, but what is truly frightening is that some human patients carry a copy of one gene specific to each strain suggesting that the virus has recombined. This is rarely seen in RNA viruses, and has never been characterized in Ebola’s broader family of filoviruses to date.

The obvious consequence of recombination capable viruses is that vaccines developed to target specific protein on the Ebola may become ineffective once the protein profile changes.

Mackenzie, Debora. “Ebola evolves deadly new trick”. New Scientist October 13-19, 2007, 12.

Proceedings of the National Academy of Sciences, DOI: 10.1073/pnas.0704076104.

IL-1 and TNF-a: Fascinating past and present

IL-1 and TNF-a: Looking to the past for answers

I recently wrote a review of the discovery of IL-1 and TNF-a for the Rheumatology Report Volume 2, Number 1 (Fall 2007). I took excerpts from this review for this blog and expanded on those I felt were most interesting to share.

For hundred of years, physicians and scientists have endeavored to discover why fever occurs and regresses in human disease. The discovery of cytokines and their inhibitors has led to a more complex understanding of the regulation and dysregulation of the immune system. The history of the discovery TNF-a and the biologic TNF-a inhibitors will be addressed as well as IL-1b and its inhibitor IL-1Ra. A review of the role of the IL-1b and TNF-a in synovial inflammation and potential targets for treatment of Rheumatoid Arthritis including gene based therapy, vaccine therapy and mesenchymal stem cell therapy will be outlined.

Coley’s toxin and the discovery of TNF-a

IN 1891, Dr. William Coley frustrated by the inability of aggressive surgery to result in a cure of sarcoma searched through the medical records of his hospital. Dr. Coley found the records of a 7 year old child who recovered from sarcoma following an infection with erysipelas. Dr Coley then proceeded to treat a patient with lymphoma with multiple injections of streptococcal cultures at 3-4 day intervals. The tumor underwent necrosis after the patient developed an attack of erysipelas and the patient remained disease free for 8 years. Throughout the remainder of his career, Dr. Coley used injection of streptococci and heat killed streptococci plus Serratia marcescens, (“Coley’s toxin”) to treat patients with malignant tumors. The dose of “Coley’s toxin” needed to produce a temperature of 40-40.5 degrees Celsius in order to induce necrosis of the tumor. Coley observed that infection resulted in a systemic response that resulted in tumor destruction.1 Many years later in 1975, an endotoxin-induced serum factor was found to be responsible for the necrosis of tumors and named Tumor Necrosis Factor. 2

Inhibition of TNF

In 1988, an inhibitory protein of TNF was isolated from the urine of febrile patients. 3 The protein inhibited with the function of TNF by blocking the binding of TNF to its’ receptor. The protein was found to bind both TNF-a and TNF-b. 4 In subsequent years, many members of the TNF-family members and their receptors were discovered as well their roles in rheumatic disease and novel ways to block their activity were developed. 5 In 1993, chimeric monoclonal antibodies to TNF-a were used to block its activity and to successfully treat rheumatoid arthritis. 6 Modification of a TNF receptor for the successful treatment of rheumatoid arthritis was accomplished in 1997 with the design of a recombinant soluble TNF receptor (p75) linked to the Fc portion of human IgG1. 7 The soluble TNF receptor (p75) binds and blocks the activity of both TNF-a and TNF-b.

The History of IL-1 and its Inhibition

In 1926, Zinsser and Tamiya discovered that contact with tissues of animals infected with tuberculosis resulted in the production of a toxic factor in uninfected cells. They suspected that a protein constituent of mycobacterial growth or of the mycobacterium itself stimulated cells to liberate a toxic substance on other cells or within the cell itself. 8 These observations may have been the first described effects of IL-1.

In the 1970s, numerous investigators simultaneously identified factors that modulate lymphocyte function including lymphocyte-activating factor, mitogenic protein, helper peak-1, t cell-replacing factor III, T cell-replacing factorMF, B cell-activating factor and B cell differentiation factor. It was determined that these factors were identical and in 1979 all were renamed IL-1. 9 The pleiotropic actions of IL-1 explain its simultaneously description and discovery by multiple investigators. In subsequent years, different receptors and inhibitors of IL-1 were discovered, importantly IL-1 Receptor Antagonist (IL-1Ra) at the University of Colorado. The first clinical observation of variations of IL-1Ra was in children with systemic juvenile inflammatory arthritis and reported in 1987. 10 In 1996, the use of recombinant human IL-1Ra, anakinra, was used to treat patients with rheumatoid arthritis. 11 More recently, IL-1Ra has been successfully used in treating patients with juvenile onset and adult onset Still’s disease. 12-14

How the synovium may be subject to inflammation in rheumatoid arthritis

The initial site of inflammation in rheumatoid arthritis has remained elusive. Ochi et al reported on the ability of fibroblastic stromal cells from the bone marrow to migrate to the joint space and form synovial tissue in the mouse model of collagen-induced arthritis. This specific population of fibroblast stromal cells can act as nurse-like cells and have the ability to interact with lymphocytes and monocytes and induce cellular differentiation and promote biological activities that mimic features of rheumatoid inflammation. 15 The findings of this study provide evidence of the bone marrow’s ability to maintain a disease state and to direct localization to the joints. It has yet to be established how the nurse-like cells of fibroblast stromal origin migrate from the bone marrow to the joint space.

A mechanism by which antibodies may gain access to joint compartment was reported by Binstadt et al, 2006. Using intravital imaging, they identified that arthritogenic antibodies from the K/BxN arthritis model caused vasopermeability localized to sites leading to the development of arthritis in normal mice. This vasopermeability was dependent upon mast cells, neutrophils and FcgRIII but not complement, TNF or IL-1. 16 Neither of these studies identified the initial site of inflammation or trigger of inflammation in rheumatoid arthritis, but provided insight into the mechanisms of continued disease activity and identify potential targets for future treatment modalities.

Cadherin- 11, an adhesion molecule, was shown to play a critical role in establishing synovial cell- to cell- contact necessary for synovial lining formation. Caherin-11 is necessary for the establishment of the K/BxN serum mediated transfer of arthritis as cadherin-11-null mice failed to develop inflammatory arthritis and had a poorly organized synovium. Similarly, use of anti-cadherin 11 in established arthritis helped to ameliorate disease in the K/BxN serum transfer mouse model of arthritis.17

IL-1b and TNF-a induce the overgrowth of synovial cells

In collagen-induced arthritis, IL-1b and to a lesser extent TNF-a were found to induce the expression of synoviolin in mouse synovial fibroblasts. IL-1b induced synoviolin transcription, which in turn enhanced IL-1b induced synovial fibroblast proliferation. The synovial fibroblasts produced more IL-1b for the induction of synoviolin leading to a positive feedback loop that may be critical in maintenance of rheumatoid inflammation. 18

Targeting inflammatory cytokines through gene based therapy

In a phase-I trial, fibroblast-like synoviocytes were transduced with a retroviral vector containing the gene for IL-1Ra and injected into the metacarpophalangeal joints. The transfer and expression of IL-1Ra was safely and successfully accomplished but no follow-up clinical studies are planned using this method. 19

A phase I dose escalation trial using delivery of a recombinant adeno-associated virus containing the TNF-receptor-Fc immunoglobulin fusion gene (tgAAC94) has been conducted and a phase I/II trial was being conducted when a study participant died.19 The woman had a low-grade fever and fatigue for several days before she received a second dose of active drug, dosed at10 trillion tgAAC94 particles per milliliter. Within a few days after the second injection, her condition worsened and she died of histoplasmosis. Last month, at the American College of Rheumatology meeting in Boston, results of an investigation into the woman’s death were released. Genetic analyses of tissue samples from the woman showed 500,000 copies/microgram of tgAAC94 in the injected knee, but fewer than 30 copies/microgram in other tissues including the other knee, liver, spleen, tonsil, and bowel. Wild-type AAV copies were found at low or undetectable levels in the various tissues. The investigation found no evidence that the agent had been contaminated with Histoplasma. 20 The FDA announced this week that the trial has been re-opened for enrollment.

Before gene therapy is successful, it needs to be determined if neutralizing antibodies to the viral vectors will render the therapy ineffective. Additionally, methods for producing vectors needs to be optimized and how therapeutic gene expression will be regulated in the human host needs to be established. 19 Methods to measure the suppression of systemic TNF activity attributable to gene therapy need to be designed to determine the activity of the gene therapy. Moreover, there are numerous safety concerns to be addressed.

TNF-a kinoid vaccination as a potential therapy for rheumatoid arthritis

Le Buanec et al, 2006 have developed a vaccine using a keyhole limpet hemocyanin-hTNF-a immunogen. When injected into hTNF-a transgenic mice in incomplete Freund’s adjuvant, a high-titer of neutralizing antibodies to hTNF-a were produced that eliminated the bioactivity of hTNF-a and resulted in reversal of arthritis. 21 Further studies are needed in other mouse models and caution should be used in the interpretation of these results, as completely limiting the function of TNF-a in humans would likely result in deleterious effects including increased risk of infection and malignancy.

Immunomodulation through the use of mesenchymal stromal cells

The report of an interdisciplinary meeting addressing the potential role for multipotent mesenchymal stromal cells in the pathogenesis and management of autoimmune diseases was published in January 2007. Mesenchymal stromal cells, derived from the bone marrow, in vitro exhibit antiproliferative effects on T and B lymphocytes, dendritic cells, natural killer cells and B cell tumor cell lines. 22,23 In the collagen-induced arthritis mouse model, injecting a murine mesenchymal cell line systemically or intra-articularly was not effective in modulating disease and labeled mesenchymal stromal cells were not found in the articular injection sites. The lack of therapeutic effect was attributed to increased levels of TNF-a and its reversal of the immunosuppressive properties of the mesenchymal stromal cells. 24 More data is needed on the optimal source of mesenchymal stromal cells as those from autoimmune diseased patients may not be equivalent to those from healthy individuals.25 Additionally, optimal timing, location and number of cells to be given have yet to be determined. 23

Final Thoughts

The extensive scientific and clinical work over the last century has led to exciting discoveries in immune system biology and the pathogenesis of autoimmune diseases. The experiences of the past with infectious agents and the responses they elicit led to the discovery of cytokines and there potential use in altering immune response to benefit individuals. Although blocking of one cytokine with current biologic therapies has led to improved management of rheumatoid arthritis, for many patients blocking only one cytokine may not be sufficient. 26 New ways of targeting cytokines and their regulation may be possible in the future with gene therapy, immunization therapies and tissue specific synovial based therapies.
We shall not cease from exploration
And the end of all our exploring
Will be to arrive where we started
And know the place for the first time.

Little Gidding V, Four Quartets, T.S. Eliot (1943)

For an interesting discussion of Dr. Coley’s work and progress made after his death see: http://www.coleytoxins.com/1893htm.

References

1. Bickels J, Kollender Y, Merinsky O, Meller I. Coley's toxin: historical perspective. Isr Med Assoc J. Jun 2002;4(6):471-472.
2. Carswell EA, Old LJ, Kassel RL, Green S, Fiore N, Williamson B. An endotoxin-induced serum factor that causes necrosis of tumors. Proc Natl Acad Sci U S A. Sep 1975;72(9):3666-3670.
3. Seckinger P, Isaaz S, Dayer JM. A human inhibitor of tumor necrosis factor alpha. J Exp Med. Apr 1 1988;167(4):1511-1516.
4. Engelmann H, Aderka D, Rubinstein M, Rotman D, Wallach D. A tumor necrosis factor-binding protein purified to homogeneity from human urine protects cells from tumor necrosis factor toxicity. J Biol Chem. Jul 15 1989;264(20):11974-11980.
5. Ashkenazi A. Targeting death and decoy receptors of the tumour-necrosis factor superfamily. Nat Rev Cancer. Jun 2002;2(6):420-430.
6. Elliott MJ, Maini RN, Feldmann M, et al. Treatment of rheumatoid arthritis with chimeric monoclonal antibodies to tumor necrosis factor alpha. Arthritis Rheum. Dec 1993;36(12):1681-1690.
7. Moreland LW, Baumgartner SW, Schiff MH, et al. Treatment of rheumatoid arthritis with a recombinant human tumor necrosis factor receptor (p75)-Fc fusion protein. N Engl J Med. Jul 17 1997;337(3):141-147.
8. Zinsser H, Tamiya T. An Experimental Analysis of Bacterial Allergy. J Ex Med. 1926;44:753-776.
9. Revised nomenclature for antigen-nonspecific T cell proliferation and helper factors. J Immunol. Dec 1979;123(6):2928-2929.
10. Prieur AM, Kaufmann MT, Griscelli C, Dayer JM. Specific interleukin-1 inhibitor in serum and urine of children with systemic juvenile chronic arthritis. Lancet. Nov 28 1987;2(8570):1240-1242.
11. Campion GV, Lebsack ME, Lookabaugh J, Gordon G, Catalano M. Dose-range and dose-frequency study of recombinant human interleukin-1 receptor antagonist in patients with rheumatoid arthritis. The IL-1Ra Arthritis Study Group. Arthritis Rheum. Jul 1996;39(7):1092-1101.
12. Fitzgerald AA, Leclercq SA, Yan A, Homik JE, Dinarello CA. Rapid responses to anakinra in patients with refractory adult-onset Still's disease. Arthritis Rheum. Jun 2005;52(6):1794-1803.
13. Vasques Godinho FM, Parreira Santos MJ, Canas da Silva J. Refractory adult onset Still's disease successfully treated with anakinra. Ann Rheum Dis. Apr 2005;64(4):647-648.
14. Aarntzen EH, van Riel PL, Barrera P. Refractory adult onset Still's disease and hypersensitivity to non-steroidal anti-inflammatory drugs and cyclo-oxygenase-2 inhibitors: are biological agents the solution? Ann Rheum Dis. Oct 2005;64(10):1523-1524.
15. Ochi T, Yoshikawa H, Toyosaki-Maeda T, Lipsky PE. Mesenchymal stromal cells. Nurse-like cells reside in the synovial tissue and bone marrow in rheumatoid arthritis. Arthritis Res Ther. 2007;9(1):201.
16. Binstadt BA, Patel PR, Alencar H, et al. Particularities of the vasculature can promote the organ specificity of autoimmune attack. Nat Immunol. Mar 2006;7(3):284-292.
17. Lee DM, Kiener HP, Agarwal SK, et al. Cadherin-11 in synovial lining formation and pathology in arthritis. Science. Feb 16 2007;315(5814):1006-1010.
18. Gao B, Calhoun K, Fang D. The proinflammatory cytokines IL-1beta and TNF-alpha induce the expression of Synoviolin, an E3 ubiquitin ligase, in mouse synovial fibroblasts via the Erk1/2-ETS1 pathway. Arthritis Res Ther. 2006;8(6):R172.
19. Adriaansen J, Vervoordeldonk MJ, Tak PP. Gene therapy as a therapeutic approach for the treatment of rheumatoid arthritis: innovative vectors and therapeutic genes. Rheumatology (Oxford). Jun 2006;45(6):656-668.
20. Gever J. www.medpagetoday.com/MeetingCoverage/ACRMeeting/tb/7390.
21. Le Buanec H, Delavallee L, Bessis N, et al. TNFalpha kinoid vaccination-induced neutralizing antibodies to TNFalpha protect mice from autologous TNFalpha-driven chronic and acute inflammation. Proc Natl Acad Sci U S A. Dec 19 2006;103(51):19442-19447.
22. Di Nicola M, Carlo-Stella C, Magni M, et al. Human bone marrow stromal cells suppress T-lymphocyte proliferation induced by cellular or nonspecific mitogenic stimuli. Blood. May 15 2002;99(10):3838-3843.
23. Tyndall A, Walker UA, Cope A, et al. Immunomodulatory properties of mesenchymal stem cells: a review based on an interdisciplinary meeting held at the Kennedy Institute of Rheumatology Division, London, UK, 31 October 2005. Arthritis Res Ther. 2007;9(1):301.
24. Djouad F, Fritz V, Apparailly F, et al. Reversal of the immunosuppressive properties of mesenchymal stem cells by tumor necrosis factor alpha in collagen-induced arthritis. Arthritis Rheum. May 2005;52(5):1595-1603.
25. Del Papa N, Quirici N, Soligo D, et al. Bone marrow endothelial progenitors are defective in systemic sclerosis. Arthritis Rheum. Aug 2006;54(8):2605-2615.
26. Dayer J. How Far Are We From Understanding Rheumatoid Arthritis? Ann Rheum Dis. 2007;66(Suppl II)(9).

HIV is a superantigen for B-cells

I have worked on a variety of human immunodeficiency virus (HIV) projects, but only recently became aware of not-so-new research suggesting the HIV envelope glycoprotein, gp 120, is an Ig superantigen for a subpopulation of B cells. A superantigen has been defined as an antigen binding 5-25% of the T or B cell population, dramatically more than the <0.01% of a conventional antigen (Goodglick, 1995).

It is well known that infection with HIV depletes CD4+ T cells. B cell repertoires, which lack CD4, are also impacted by the virus (Berberian, 1993). Antibody specificity is determined by the variable regions of their heavy (VH) and light chains. The VH genes are divided into 7 families (VH1-7) that are distinguished by at least 80% nucleotide homology in certain VH framework regions. In adult peripheral B cells, VH3 comprises about half of the expressed VH repertoire (Scamurra, 2000). HIV gp120 stimulates endogenous B cells from the VH3 family by binding to membrane Ig, outside the conventional, (hypervariable) Ag-binding site. HIV-1 gp120 has thus been labeled a superantigen, binding an estimated 4-6% of the B cell population (Goodglick, 1995). This binding results in an increase in VH3 B cells in early disease. This overactive humoral state is followed by a decline in B cell response which is prior to the significant depletion of CD4+ T cells (Karray, 1997). The resulting B cell loss and/or shift in distribution may result in impaired humoral responses to infections and vaccines (Scamura, 2000).

After writing the above, I received some pre-blog feedback from Elisabeth Bowers, a pre-doctoral fellow in the Microbiology department working in this area. She noted that a direct association between the increase in VH3 B cells in early disease and the B cell dysfunction seen during infection has not been shown. (B cell dysfunction includes hypergammaglobulinemia –a high amount of antibody in the blood, lymphadenopathy, and increased risk of B cell lymphomas.) The B cell dysfunction may be, in part, a result of increased VH3 B cells, but much of the response is directed against HIV. “So the current hypothesis for some (but not all) of the B cell dysfunction is humoral immune reaction to the virus itself, leading to B cells producing virus-specific antibodies, and ‘bystander’ B cell responses – B cells that are not specific to virus, but still become activated anyway (these may not all be VH3 B cells) (Shirai, 1992; Amadori, 1990; Schnittman, 1986). B cell abnormalities have been directly associated with high viral loads (Viau, 2007).”

Berberian, et al., “Immunolglobulin VH3 gene products: natural ligands for HIV gp120”. Science. 1993. 261: 1588-91.

Goodglick, et al,. “Mapping the Ig superantigen-binding sit of HIV gp120”. The Journal of Immunology. 1995. 155: 5151-59.

Karray, et. al., “Identification of the B cell superantigen-binding site of HIV-1 gp120”. PNAS. 94: 1356-60.

Scamurra et al., “Impact of HIV-1 infection of Vh3 gene repertoire of naïve human B cells”. The Journal of Immunology. 2000. 164: 5482-91.

Shirai et al., “Human Immunodeficiency Virus Infection Induces Both Polyclonal and Virus-specific B Cell Activation”. The Journal of Clinical Investigation. 1992. 89: 561-566.

Amadori et al., “B-cell activation and HIV-1 infection: deeds and misdeeds”. Immunology Today. 1990. 11(10): 374-379.

Schnittman et al., “Direct Polyclonal Activation of Human B Lymphocytes by the Acquired Immune Deficiency Syndrome Virus”. Science. 1986. 233: 1084-1086.

Viau et al., “Direct impact of inactivated HIV-1 virions on B lymphocyte subsets”. Molecular Immunology. 2007. 44: 2124-2134.

Possible reasons why the Merck HIV vaccine failed

Going along with the theme of the previous few blog posts, I was going to blog about the failed Merck HIV vaccine trials.  Instead, I decided to delve a little bit deeper into the hypotheses attempting to explain WHY this vaccine, even the promising results of increased CD8+ killer T cell numbers, may have ultimately failed.

The MRKAd5 vaccine was based on the premise of inserting copies of 3 artificially generated HIV genes into a genetically engineering viral vector (2).  The vector used in the HIV vaccine, and many other viral vaccines, is adenovirus5, an inactivated form of a "common cold" virus.  The hopes for this vaccine were that the immune system would recognize the viral components enough that if a vaccinated individual were exposed to HIV, there would be enough of a response to prevent, or at least delay, the onset of AIDS (1).  However, after a conference in Seattle to discuss the new evidence that the MRKAd5 vaccine is "not efficacious" and may result in INCREASED risk for HIV infection (3), several ideas have been brought forth in an attempt to explain why the vaccine failed.  One theory is based on the idea that if a vaccinated volunteer was previously exposed to adenovirus5 (the viral backbone of the vaccine), the subsequent response would be biased towards CD4+ cells rather than the Cd8+ cells the vaccine was intended to amplify.  An increase in CD4+ cells only results in the production of more cells that the HIV virus can go on to infect (Dr. Keith Gottesdiener, unpublished data). Some preliminary results suggest that some people who have prior immunity to adenovirus5 may indeed be the ones who are at higher risk for HIV infection.  The second theory depends on the idea that perhaps this system generates too much antigen for a prolonged period of time.  The excess antigen causes "exhaustion" of the immune cells, which then fail to proliferate and die (6).

Regardless of the actual unforeseen mechanisms that caused the MRKAd5 vaccine to fail,  the outcome of this trial raises questions concerning the use of similar adenovirus vectors in vaccine creation.  Several other vaccine trials, including an Ebola vaccine and another HIV vaccine, have been slowed, postponed or altered so that the test population pool are individuals with low risk of exposure to colds (4).  It also reminds us that while data generated in model organisms is invaluable, care must be taken when extrapolating the putative animal model into human applications, as we've seen several times during this course.  In the case of the MRKAd5 vaccine, the data generated in primates suggested that the vaccine upregulated the number of CD8+ cells (7), although later on, it was shown in mouse that these additional cells may not be completely functional (2).

1.  Ho, DD.  (2007).  A Shot in the Arm for AIDS Vaccine Research.  PLoS Med 2(2) :e36.

2.  Lin, SW et al.  (2007).  Recombinant adeno-associated virus vectors induce functionally impaired transgene product-specific CD8 T cells in mice.  J Clin Invest.  2007 Nov. 15 [Epub ahead of print].

3.  Cohen, J.  (2007).  Did Merck's vaccine cause harm?  Science 312(5853):1048-9.

4.  Stark, K.  (2007).  HIV trial's halt reverberates.  www.philly.com/philly/business/20071116_HIV_trials_halt_reverberates.html

5.  Cohen, J.  (2007).  Promising AIDS vaccine's failure leaves field reeling.  Science 318(5847):28.

6.  Kaiser Daily AIDS/HIV report from November 16, 2007.  www.kaisernetwork.org/daily_reports/rep_index.cfm?DR_ID=48904

7.  Shiver, JW and Emini EA.  (2004).  Recent advances in the development of HIV-1 vaccines using replication-incompetent adenovirus vectors.  Annu Rev Med 55:355-72.

Believe them or not, interesting associations among “old friends”, gut dysfunction and autism

Ever since our first Immunology class, the “old friends”, commensal flora, had caught my attention since they seemed to be able to survive from human immune system’s attack and I was very interested in finding out how they do that. It turned out that this is still a largely unknown question. People agree that the establishment of commensal flora after birth represents a major developmental and functional stimulus to the immune system. The fighting between the immune system and the commensal flora finally creates a mucosal immune homeostasis (1). In addition to stimulate immune system to mature, the “old friends” also support digestion, absorption and metabolism; they compete with pathogens in our intestine for space and resources too (2).

The most interesting thing I found during my web search about commensal flora is a hypothesis on the association between autism and gut dysfunction which could be caused by the altered “old friends”. Autism is a developmental disorder for which there is no drug or even a known cause. One of the hypotheses of its cause is the alteration of gut flora leads to increases in intestinal permeability, also known as “leaky gut” which allows the passage of neuroactive peptides digested from food origin into the blood and then into the cerebrospinal fluid to interfere directly with the function of the CNS (3). In the presence of an alteration of the gut flora, or immune dysregulation, or agenetic predisposition, there is a sustained chronic inflammation and release of calprotectin from neutrophils. The inflammatory response of the intestinal epithelial cells to pathogeneic and altered gut flora can lead to increases in intestinal permeability thus allow the passage of toxins, microbes, undigested food, waste or larger than normal macromolecules (2). Digestion products of natural foods such as cow’s milk and bread are able to enter the blood through the leaky mucosa and induce antigenic responses, as well as interfere directly with the central nervous system. Casomorphins and gliadomorphins are good examples as the short chain neuroactive peptides digested from dietary gluten and casein in the lumen of the small intestine by the action of pancreatic and intestinal peptidases. These peptides, which are structurally similar to endorphins, are called exorphins to reflect their dietary origin (3). Interestingly, a substantial group of autistic children have been diagnosed to have increased intestinal permeability (4, 5).

It seems that the true, unaltered “old friends” are really versatile good friends.


Reference:
1. http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=eurekah.section.
2. http://littlecanaries.org/gastro.htm
3. White, JF. Intestinal Pathophysiology in Autism Exp Biol Med (Maywood) 2003, 228, 639–649.
4. Horvath K, Zielke RH, Collins J, et al. Secretin improves intestinal permeability in autistic children. J Pediatr Gastroenterol Nutr 2000;31: S30-31.
5. D'Eufemia P, Celli M, Finocchiaro R, et al. Abnormal intestinal permeability in children with autism. Acta Paediatr 1996;85: 1076-1079.