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A report of two cases and systematic literature review. World J Gastrointest Surg . 2013;5(3):37–42. 48. F ollmar KE, Condron SA, Turner II, Nathan JD, Ludwig KA. Treatment of metronida -
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children: A comprehensive review. Curr Med Res Opin . 2013;29(8):967–984. 50. W ilkins TD, Lyerly DM. Clostridium difficile  testing: After 20 years, still challenging. J Clin Microbiol. 2003;41:531–534. 51. K yne L, Hamel M, Polavaram R, Kelly C.  Health care costs and mortality asso -
ciated with nosocomial diarrhea due to Clostridium difficile. Clin Infect Dis . 2002;34:346–353. 52. Qiu B, Pothoulakis C, Castagliuolo I, Nikulasson S, LaMont JT . Participation of reac -
tive oxygen metabolites in Clostridium difficile  toxin A-induced enteritis in rats. Am 
J Physiol . 1999;276(2 Pt 1):G485–G490. 53. McK enzie SJ, Baker M, Buffinton G, Doe W. Evidence of oxidant-induced injury to 
epithelial cells during inflammatory bowel disease. J Clin Invest . 1996;98:136–141. 54. T riadafilopoulos G, Pothoulakis C, Weiss R, Giampaolo C, LaMont J. Comparative 
study of Clostridium difficile toxin A and cholera toxin in rabbit ileum. Role of prosta -
glandins and leukotrienes. Gastroenterology . 1987;92:1174–1180. 55. Studahl M, Lindquist L, Eriksson BM, Günther G, Bengner M, Franzen-Röhl  E, 
Fohlman J, Bergström T, Aurelius E. Acute viral infections of the central ner -
vous system in immunocompetent adults: Diagnosis and management. Drugs . 2013;73(2):131–158. 56. Field HJ, Vere Hodge RA. Recent developments in anti-herpesvirus drugs. Brit Med 
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Acad Sci . January 27, 1975;243:395–402.114 Dimethyl Sulfoxide (DMSO) in Trauma and Disease
 58. MacCallum FO, Juel-Jensen BE. Herpes simple x virus skin infection in man treated 
with idoxuridine in dimethyl sulphoxide. Results of double-blind controlled trial. Brit 
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McKeough MB, Rowe NH. Early application of topical 15% idoxuridine in dimethyl 
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L, Ledo A, Mascaró JM. A topical solution of 40% idoxuridine in dimethyl sulfoxide compared to oral acyclovir in the treatment of herpes zoster. A double-blind multicenter clinical trial. Med Clin  (Barc ). 1992;98(7):245–249. 61. Freeman DJ, Sheth NV , Spruance SL. Failure of topical acyclovir in ointment to pen -
etrate human skin. Antimicrob Agents Chemother . 1986;29(5):730–732. 62. Aguilar JS, Ro y D, Ghazal P, Wagner EK. Dimethyl sulfoxide blocks herpes simplex 
virus-1 productive infection in vitro acting at different stages with positive cooperativity. Application of micro-array analysis. BMC Infect Dis . 24, 2002;2:9. 63. Whitle y RJ, Soong SJ, Dolin R, Galasso GJ, Chien LT, Alford CA. Adenine arabinoside 
therapy of biopsy-proved herpes simplex encephalitis. National Institute of Allergy and Infectious Diseases collaborative antiviral study. N Engl J Med . 1977;297:289–294. 64. Henninger R W, Reed SM, Saville WJ, Allen GP, Hass GF, Kohn CW, Sofaly C. Outbreak 
of neurologic disease caused by equine herpesvirus-1 at a university equestrian center. J Vet Intern Med . 2007;21(1):157–165. 65. W ong D, Scarratt WK. Equine herpes myeloencephalopathy in a 12-year-old American 
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of antifungal susceptibility variables for a semiautomated methodology. J Clin Microbiol . 2001;39(7):2513–2517. 72. Hazen KC. Influence of DMSO on antifung al activity during susceptibility testing in 
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6DMSO in Clinical 
Microbiology
HOW BACTERIA ACHIEVE ANTIBIOTIC RESISTANCE
Antibiotic resistance by bacteria refers to the ability of microorganisms to withstand 
the killing effects of one or more antibiotics. Bacteria resistant to multiple antibiotics are considered multidrug resistant and are referred more colloquially to as superbugs. Drug-resistant bacteria pose one of the most serious and growing threats to 
health and survival and continue to be a global concern to contemporary medicine. Antibiotic-resistant infections complicate treatment and increase morbidity and mortality. These infections are difficult to treat with first-line antibiotics and often result in 
second-line treatments that promote longer-lasting illnesses, more doctor visits or extended hospital stays, and the need for more expensive and less safe medications. Some resistant infections can cause death. The first step in the emergence of antibiotic-resistant microorganisms is a genetic 
change in a bacterium. There are two ways that can happen. The first way is by spon -
taneous mutation of the bacterium’s DNA. Many antibiotics kill bacteria by inacti -
vating an essential bacterial protein, but either the microbe can develop a mutation that either prevents binding of the antibiotic to the protein or can increase the pro -
duction of that protein. Last, the bacterium can shed the targeted protein from its cell body. The second way bacteria can become resistant to antibiotics is by the transfer of immunity from a resistant bacterium to its progeny, thus ensuring the survival of subsequent generations of microbes. In some countries, including the United States, patients expect and demand anti -
biotics from doctors, even in situations where they are inappropriate or ineffective. Because microbes are always mutating, some random mutation eventually will 
 protect against the drug. As people enjoy the benefits of antibiotics, their misuse 
promotes antibiotic resistance in many lethal bacteria. Resensitizing resistant bacteria to antibiotics so that they can be killed is the 
objective of searching for antimicrobial adjuvants that can be used to boost otherwise ineffective antibiotics against methicillin-resistant Staphylococcus aureus  (MRSA), 
Mycobacterium tuberculosis , Pseudomonas , Clostridium difficile , and other super -
bugs that can cause lethal infectious outbreaks in hospitals and institutions. The emergence of multidrug-resistant and extensively drug-resistant M. tuber -
culosis  throughout the developing world is very disturbing in the present scenario 
of tuberculosis (TB) management. TB has always been a global problem and tends to increase in prevalence mainly in the developing countries of the world, 116 Dimethyl Sulfoxide (DMSO) in Trauma and Disease
many of the affected living in poverty, but is also problematic in industrialized 
nations, including the United States.1
M. tuberculosis  usually attacks the lungs, but these pathogens can invade any part 
of the body such as the kidney, spine, and brain. There are 10 drugs currently approved by the U.S. Food and Drug Administration  
(FDA) for treating TB. Before the emergence of antibiotic-resistant pathogens, potent drugs such as rifampicin, isoniazid, ethambutol, and pyrazinamide were effective as first-line anti-TB agents in the treatment of latent or active TB. Most patients with active TB can be treated initially with isoniazid, rifampicin, pyrazinamide, and eth-ambutol for 8 weeks, and this regimen can be followed by 18 weeks of treatment with isoniazid and rifampicin if needed. 2
Multidrug-resistant TB is defined by resistance to the two most effective first-line 
drugs, isoniazid and rifampicin.3 When bacteria are resistant to second-line antibi -
otics, they are called extremely resistant. Recently, strains of M. tuberculosis  have 
been identified that are resistant to all available antibiotics. This is representative of the major problems of antibiotic resistance and the cause of considerable concern if these strains begin to quickly spread worldwide. Approximately 3.3% of new multidrug-resistant TB cases are resistant to both 
isoniazid and rifampicin, so other drug combinations must be tried. However, most of these drug combinations fail to fully treat TB, and consequently, there is a fun -
damental need to explore alternative anti-TB agents that can kill strains resistant to currently available drugs and for new drugs that are better tolerated and will shorten treatment regimens. 4
DMSO IN MULTIDRUG-RESISTANT TUBERCULOSIS
Researchers working in the field of microbiological pharmacotherapeutics are looking to find ways to make first-line antibiotic therapies effective against drug-resistant strains of bacteria. Part of this research is to search for adjunct therapy that can help current antibiotics function more effectively, even in multidrug-resistant strains of bacteria. Many of the early studies examining the role of dimethyl sulfoxide (DMSO) in 
multidrug-resistant TB were done by Russian and Polish scientists in the mid-1970s. A summary of these studies details some of the properties observed when DMSO was used as an adjuvant to antibiotics against M. tuberculosis –resistant bacteria or by itself. In one study, isoniazid-resistant TB bacteria were inoculated into guinea pigs that 
were divided into two groups: the first group underwent isoniazid treatment and the second group was treated with a single oral dose of DMSO and isoniazid 2 weeks after inoculation with tubercle bacilli. 5 When control animals died from the bacterial 
inoculation after 80 days, samples were taken from liver, lymph nodes, and spleens and cultured and examined microscopically. 5 All guinea pigs treated with DMSO–  
isoniazid survived beyond 1 year. Animals not treated with DMSO yielded tubercle bacilli in the collected organs and appeared entirely resistant to isoniazid treatment. 5 
These findings indicated that reversion to isoniazid sensitivity by DMSO permitted a cure of the experimental animals infected with an isoniazid-resistant strain of tuber -
cle bacilli, suggesting that this drug combination may be clinically useful in human 
 antibiotic-resistant TB.117 DMSO in Clinical Microbiology
A subsequent study by the same investigators replicated the guinea pig experiments 
in vitro  by isolating isoniazid- and rifampicin-resistant tubercle bacilli from patients 
with pulmonary TB.6 This study consisted of cultures containing 61 strains of tubercle 
bacilli resistant to isoniazid and 19 strains resistant to rifampicin. DMSO 5% exposure 
to the cultures resulted in 19/61 strains resistant to isoniazid reverted to sensitivity and all 19 rifampicin-resistant strains also reverted to sensitivity after DMSO exposure. 6 
The data showed that complete reversion of rifampicin sensitivity was obtained with DMSO and also reversion of some isoniazid-resistant tubercle bacilli strains indicated support for a clinical study of these antibiotics combined with DMSO. DMSO has also been reported to stabilize rifampicin for periods of up to 8 months 
without any loss of potency by this antibiotic. 7
Aside from its preservation of shelf life for many drugs, a common use for DMSO 
is its ability to act as a safe and versatile solvent for many antibiotics, including those used for antibiotic-resistant pathogens. 8 There are no data relating to DMSO acting 
or synergistically potentiating antibiotic activity when it is used as a solvent. These findings created much interest among investigators interested in antibiotic-
resistant therapy, and reports soon appeared in an attempt to further examine the ability of DMSO to restore sensitivity to chemotherapeutic compounds used to treat various resistant bacterial strains. M. tuberculosis  strains, resistant to rifampicin, isoniazid, streptomycin, 
paraminosalicylic acid, thioacetozone, ethambutol, and prothionamide, were grown in cultures containing these drugs together with either DMSO at 1% concentration or water. 9 Susceptibility tests were applied, and resensibilization 
to ethambutol, streptomycin, and thioacetozone was observed only in DMSO-treated cultures. 9 Moreover, many M. tuberculosis  strains resistant to isoniazid 
and all strains resistant to rifampicin became sensitive to these drugs when com -
bined with DMSO.9
The ribosomal subunit 30S is considered a site of action of streptomycin and also 
a site of resistance for certain strains of Escherichia coli  and other bacteria. These 
mutant germs show an impermeability to streptomycin that confers resistance to the drug’s action. 10 The use of DMSO to overcome E. coli  resistance to streptomycin in 
cultures showed that these pathogens decreased their permeability and increased their uptake of streptomycin, thus allowing a killing effect by the antiobiotic. 11 The 
effect by DMSO on E. coli  cell membrane with respect to enhanced streptomycin 
uptake can only be surmised. Drug-resistant isolates of Mycobacterium avium–Mycobacterium intracellulare  
were isolated and cultured in vitro  in the presence of single drugs alone and in com -
bination with 2.5% DMSO.12 Bacterial isolates were exposed to first-line antibiotics 
used for the treatment of TB, including rifampicin, streptomycin, ethambutol, and isoniazid, and all showed a degree of killing power that varied depending on the bac-terial strain, but with the addition of DMSO to each antibiotic, a 26%–30% increase in inhibitory bacterial growth was observed. 12
Thirty-two patients with destructive pulmonary TB were treated with strep -
tomycin and penicillin in 10% and 25% solutions of DMSO through inhala -
tion. Fourteen of the patients showed absence of M. tuberculosis excretion. Most patients showed improvement, including a decrease in manifestations of 118 Dimethyl Sulfoxide (DMSO) in Trauma and Disease
nonspecific endobronchitis, a decrease in the perifocal infiltration, and a decrease 
in the dimensions of destruction of lung tissue.12a
These early studies showing the ability of DMSO to enhance antibiotic sensitivity 
to drug-resistant pathogens partly introduced the concept of the adjuvant, defined as a compound that could increase the killing power of antibiotics and antimicrobial agents that by themselves failed to inhibit bacterial growth in vitro  or in vivo . No longer would magic bullets like penicillin create their magic on a growing list 
of superbugs; now, it was necessary to find adjuvants that could help fight germs and postpone the day when first-line antibiotics would no longer work. The superbugs have devised ingenious ways to evolve in order to elude and weaken 
antibiotic drug action. The first of these is to increase their membrane permeability, thus preventing the antibiotic to accumulate to toxic levels within the cell. A second way is to make the drug less toxic using special enzymes to reduce drug potency, a third way is to inactivate the antibiotic, and a final way is to follow a metabolic pathway not targeted by the drug. 13
One strategy to get around these neutralizing techniques used by superbugs on 
antibiotic monotherapy is to combine two to four different antibiotics in a shot-gun approach. However, the downside produced by multiple drug administration is seri -
ous side-effects and the real possibility of generating more multidrug-resistant bacte -
ria, a therapeutic nightmare. Adjuvants to antibiotics may be more useful if they can either target the bacterial membrane, be more toxic to pathogens, or follow bacte -
rial pathways that can be targeted by antibiotics. This is where DMSO, and similar agents, may become clinically useful, especially in drug-resistant TB. Globally, an estimated 8.7 million people fell ill due to TB and 1.4 million died 
from this disease in 2011. TB is second only to HIV/AIDS as the greatest killer worldwide due to a single 
infectious agent. Thus, with almost 9 million new cases each year, TB remains one of the most 
feared diseases on the planet. Despite a reduction in incidence in several regions of the planet, TB remains a pandemic of worldwide significance. There is an urgent need to develop better regimens, to shorten treatment, and to effectively manage both drug-sensitive and drug-resistant disease. TB is spread from person to person through the air. When people with lung TB 
cough, sneeze, or spit, they propel the TB germs into the air. A person needs to inhale only a few of these germs to become infected. When a person develops active TB, the symptoms they develop, cough, fever, 
night sweats, weight loss, chest pains, and weakness, may be mild for many months. This mild beginning of TB can lead to delays in seeking care and may result in 
 transmission of the Mycobacterium  to others. It is estimated that persons who develop TB can infect a dozen other people 
through close contact over the course of a year. Without proper and immediate treat -
ment, up to two-thirds of people with TB will die. Multidrug-resistant TB is a type of TB caused by mycobacteria that do not 
respond to isoniazid and rifampicin, the two most powerful, first-line, and standard anti-TB drugs. When bacteria fail to respond to first-line antibiotics, second-line antibiotics must be used.119 DMSO in Clinical Microbiology
As bad as drug-resistant TB can become, an even worse outcome is the develop -
ment of more severe drug resistance. This severe resistance to antibiotics can lead to 
extensively drug-resistant TB, a form of multidrug-resistant TB that responds to even fewer available medicines, including the most effective second-line anti-TB drugs. Multidrug-resistant M. tuberculosis  and extensively drug-resistant TB are rec -
ognized as a one of the most serious challenges to global health with no solution in sight. 14
A strategy to alter cell membrane permeability to overcome multidrug-  r esistant 
M.  tuberculosis  strains, using a combination of DMSO at varying doses and 
ethambutol, was explored by Jagannath and colleagues.15 This group was aware of 
DMSO’s ability to penetrate biological membranes16 and reasoned that combining it 
with a membrane detergent such as ethambutol would enhance the delivery of anti-  
tuberculous drugs at subminimum inhibitory concentrations.15
M. tuberculosis  bacilli contain an unusual lipid-rich outer layer, containing 
mycolic acids, a unique characteristic among Gram-positive bacteria. This powerful lipid-rich layer in M. tuberculosis  represents a formidable barrier for antibiotics to 
penetrate, although the synthesis of its cell wall has become a target of several effec -
tive antibiotics, such as isoniazid and ethambutol, which presumably interfere with cell wall mycolic acid production. In the study by Jagannath and colleagues,
15 three multiple-drug resistant strains 
of M. tuberculosis  resistant to rifampicin and ethambutol were suspended in broth 
and incubated for 3–5 days with ethambutol and 2%–5% DMSO. Bacilli were then washed to remove traces of ethambutol–DMSO and exposed to isoniazid, streptomy -
cin, and rifampicin. It was found that combining DMSO with ethambutol resulted in a 16–64-fold increase for rifampicin inhibition of M. tuberculosis  strains, a 16–33-fold 
increase for streptomycin, and a 4–16-fold increase of sensitivity to isoniazid. 15
Moreover, DMSO or streptomycin by themselves showed no significant inhi -
bition of M. tuberculosis  when compared to untreated controls, but combining 
DMSO–streptomycin completely eliminated the cultured bacilli after 7 days incu-bation. 15 This study indicated that the action of DMSO combined with the effect of 
ethambutol had a significant effect on delaying or damaging cell wall biosynthesis in M.  tuberculosis multidrug-resistant strains and may have altered the bacilli’s 
cell wall permeability to enhance antibiotic delivery of rifampicin, isoniazid, and streptomycin (Figure 6.1). 15
This experiment has not been replicated in vivo , despite its clinical implications 
in drug-resistant TB. Another possibility that has not been explored as far as DMSO’s adjuvant action 
to make multidrug-resistant M. tuberculosis  bacilli more sensitive to antibiotic treat -
ment is the effect of DMSO on inner membrane protein synthesis associated with energy metabolism in microorganisms such as E. coli . 17 All cells are bound by a cell 
membrane, but although the membranes of all cells are quite similar, those of bacte -
ria differ from eukaryotic cells. Bacterial cell membrane differences allow for selective action of antimicrobial 
agents. This little-studied action by DMSO on bacterial proteins responsible for energy metabolism would classify DMSO as a bacterial antimetabolite. Bacterial antimetabolites function in two ways: (1) by competitive inhibition of bacterial 120 Dimethyl Sulfoxide (DMSO) in Trauma and Disease
enzymes and (2) by erroneous incorporation into bacterial nucleic acids, where 
proper base pairs cannot form during DNA replication and transcription (Figure 6.1). It is unlikely that DMSO would act on bacterial cell membrane disruption by the 
second mechanism, since it lacks a structure and stereochemistry similar to purine and pyrimidine nucleotides found in microorganisms. This is not the case with some antibiotics, which have a similar chemical structure to purine and pyrimi -
dines in microorganisms and are able to disrupt these pathogens by damaging their DNA. 18
The mechanism involved in DMSO’s ability to alter bacterial cell membrane 
remains unclear although it is known that DMSO affects the permeability of the cell membrane to facilitate the uptake of DNA,
19 and it is reported that DMSO destroys 
or damages the integrity of various bacterial membranes by increasing their mem -
brane permeability.20
DMSO may act to facilitate transfection by opening transient pores or holes in the 
bacterial cell membrane, a process that would allow increased uptake of antibiotics or nucleic acids isolated from bacteriophages, which are harmful to bacteria and are otherwise kept from entry into the inner bacterial membrane (Figure 6.1). 19
Recently, some support for this concept gained ground when it was reported that 
DMSO can induce water pores in biological membranes, which in turn promotes the transport of active substances across cell membranes. Such membrane transport activity by DMSO could have significant clinical appli -
cation for multidrug-resistant microorganisms.Action by DMSO in bacterial resistance to antibiotics
Protein synthesis inhibition by DMSO
Transcription Translation
Protein
Inhibition of ener gy metab olite s
Mycobacte rium tu bercu losismRNACell wall synthesis inhibition by DMSO
FIGURE 6.1  Potential inhibition of multidrug-resistant M. tuberculosis  by DMSO when 
used as an antibiotic adjuvant, based on in vitro  studies.15,17,19–21 DMSO may act to delay or 
damage cell wall biosynthesis, thus allowing a greater concentration of first-line antibiotic penetration of the bacterial cell, a process that can alter transcription of mRNA and transla -
tion of protein synthesis impacting on the production of energy metabolites needed for bacte -
rial cell growth and function.121 DMSO in Clinical Microbiology
In summary, DMSO has two important qualities that can facilitate the treatment 
of antibiotic-resistant organisms. The first is its potential as a powerful and safe sol -
vent of many water-insoluble compounds; this ability can diminish the final toxicity 
of the antibiotic or antibacterial compound. The second quality shown by DMSO is its ability to potentiate the penetration of 
antibacterial drugs by either punching a hole  in the invading organism’s cell wall,15,21 
for example, that of M. tuberculosis , to enhance antibiotic drug penetration and 
concentration inside the cell wall. Another mechanism by DMSO may involve its inhibition of the growth of pathogens through its blocking action on the pathogen’s protein synthesis, which can impact the reduced production of the organism’s energy metabolites and pathogenic potential. REFERENCES
 1. Hyman CL. Tuberculosis: A survey and review of current literature. Curr Opin Pulm 
Med. 1995;1(3):234–242. 2. Potter B, Rindfleisch K, Kraus CK. Management of active tuberculosis. Am Fam Phys. 2005;72(11):2225–2232. 3. Kalokhe AS, Shafiq M, Lee JC, Ray SM, Wang YF, Metchock B, Anderson AM, Nguyen 
ML. Multidrug-resistant tuberculosis drug susceptibility and molecular diagnostic test -
ing. Am J Med  Sci. 2013;345(2):143–148. 4. Field SK, Fisher D, Jarand JM, Co wie RL. New treatment options for multidrug-  resistant 
tuberculosis. Ther Adv Respir Dis . 2012;6(5):255–268. 5. Szydlo wska T, Pawlowska I. In vivo studies on reversion to sensitivity of INH-resistant 
tubercle bacilli under the influence of dimethylsulfoxide (DMSO). Arch Immunol Ther 
Exp (Warsz ). 1974;22(4):559–561. 6. Szydło wska T, Pawłowska I. Comparative studies on the influence of dimethylsulfoxide 
(DMSO) on reversion to sensitivity to isonicotinic acid hydrazide (INH) and rifam -
picin (RMP) in resistant strains of tubercle bacilli. Arch Immunol Ther Exp  (Warsz ). 1976;24(4):575–577. 7. Karlson AG, Ulrich JA. Stability of rifampin in dimethylsulfoxide. Appl Microbiol . 1969;18(4):692–693. 8 . Ollinger J, Baile y MA, Moraski GC, Casey A, Florio S, Alling T, Miller MJ, Parish T. 
A dual read-out assay to evaluate the potency of compounds active against Mycobacterium 
tuberculosis . PLoS ONE . 2013;8(4):e60531. 9. Müller U, Urbanczik R. Influence of dimeth yl sulfoxide (DMSO) on restoring sensi -
tivity of mycobacterial strains resistant to chemotherapeutic compounds. J Antimicrob 
Chemother. 1979;5(3):326–327. 10. Anand N, Da vis BD, Armitage A. Uptake of streptomycin by Escherichia coli . Nature . 1960;185:23–24. 11. Feldman WE, Punch JD, Holden PC. In vivo and in vitro effects of dimethyl sulf -
oxide on streptomycin-sensitive and -resistant Escherichia coli . Ann NY Acad Sci . 1975;243:269–277. 12. Nash DR, Steingrube V A. In vitro drug sensitivity of M. avium –intracellulare  complex 
in the presence and absence of dimethyl sulphoxide. Microbios . 1982;35(140):71–78. 12a. Liubinets VI, Kruk MV . Dimexide in the treatment of endobronchitis in patients with 
destructive forms of pulmonary tuberculosis. Zh Ushn Nos Gorl Bolezn . November–
December 1969;29(6):68–71. 13. Alekshun MN, Le vy SB. Molecular mechanisms of antibacterial multidrug resistance. Cell. 2007;128:1037–1050.122 Dimethyl Sulfoxide (DMSO) in Trauma and Disease
 14. W orthington RJ, Melander C. Combination approaches to combat multidrug-resistant 
bacteria. Trends Biotechnol . 2013;31(3):177–184. 15. Jag annath C, Reddy VM, Gangadharam PR. Enhancement of drug susceptibility of 
multi-drug resistant strains of Mycobacterium tuberculosis  by ethambutol and dimethyl 
sulphoxide. J Antimicrob Chemother . 1995;35(3):381–390. 16. Narula PN. The comparative penetrant-carrier action of dimethyl sulfoxide and ethyl 
alcohol in vivo. Ann NY Acad Sci . 1967;141(1):277–278. 17. Sk otnicki ML, Rolfe BG. Effect of dimethyl sulphoxide on the expression of nitrogen 
fixation in bacteria. Aust J Biol Sci . 1977;30(1–2):141–153. 18. Harle y JP, Klein DA, Prescott LM. Microbiology. Wm. C. Brown Publishers, Dubuque, 
IA, 1993. 19. Melk onyan H, Sorg C, Klempt M. Electroporation efficiency in mammalian cells is 
increased by dimethyl sulfoxide (DMSO). Nucleic Acids Res . 1996;24(21):4356–4357. 20. Murata Y , Watanabe T, Sato M, Momose Y , Nakahara T, Oka S, Iwahashi H. Dimethyl 
sulfoxide exposure facilitates phospholipid biosynthesis and cellular membrane prolif -
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ide (DMSO) induced transient water pores in cell membranes. Mol Membr Biol . 2012;29(3–4):107–113.123
7DMSO in Malignancy
DMSO AND NEOPLASIA
Neoplasia is characterized by populations of precursor cells with the ability to grow 
or divide abnormally. This growth of cells exceeds those of normal tissues around them, which can persist in the same excessive manner even when the promoting stimulus has dissipated. The abnormal growth can create a tumor or mass that can be classified as benign, 
premalignant, or malignant and can also result in nontumor proliferation of imma -
ture white blood cells, such as those seen in leukemia. 1 The malignant neoplastic 
cells primarily result from damage to DNA, which originates from a variety of endogenous (inside the body) or exogenous (outside the body) sources. 2 Whatever 
the source of the stimulus, there is a deficiency in the DNA repair damage system, which can lead to DNA mutations and progression of the malignant process. 2
An enhanced understanding in the past 50 years of the transformational process 
of neoplastic cancer cells that mediate their recruitment, activation, programming, and persistence presents interesting new targets for anticancer therapy. 3
Cryoprotective agents such as dimethyl sulfoxide (DMSO) are reported to induce 
erythroid differentiation of Friend leukemic cells in vitro .4 Since malignant cells are 
known to have the capacity to differentiate into benign cells,5 chemical agents that 
can induce such cell differentiation are frequently associated with a decrease in, or a loss of, malignant potential. 5
The complex problem of how to study and treat leukemia took a giant step for -
ward in 1966, when a virus-induced erythroleukemic cell line in mice was estab -
lished in the laboratories of Friend et al.6
FRIEND LEUKEMIA CELLS
The observations obtained in subsequent years using Friend’s virus-induced eryth -
roleukemic cell line suggested that it might be possible to treat malignant disease by inducing the differentiation of the tumor cells in vivo . 7,8 The Friend leukemia system 
provided an excellent model for testing this concept. Friend leukemia cells (FLCs) can be induced to differentiate in vitro  by any one of a number of cryoprotective 
compounds, DMSO being the most notable of that group. 9 This FLC line has pro -
vided a mother lode of experiments in cancer research and is still in use today in the study of cell differentiation. When FLC is inoculated intravenously into mice, malignant disease develops 
from leukemic cell infiltration of the spleen, bone marrow, liver, and lymph nodes. 9 
When FLC is inoculated locally by subcutaneous injection into mice, the cells 124 Dimethyl Sulfoxide (DMSO) in Trauma and Disease
develop into a tumor, which has the characteristics of a myeloblastoma.9 Most of 
these animals die when the tumor spreads to other tissues. DMSO has been shown to be a prototypical inducer of cell differentiation whose 
activity was discovered when it was exposed to FLC, providing the first example of a 
nonphysiological chemical agent that possessed the ability to reverse the differentia -
tion block imposed in leukemia cells.10
Studies in the 1970s indicated that induced differentiation found in vitro  by dif -
ferentiating agents, including DMSO, could have potential to the treatment of malig -
nant disease in vivo . To prove this point, many experiments were designed using FLC 
and DMSO to see whether malignant cell differentiation could be turned benign.10 
Thus, the hunt for an ideal chemotherapeutic agent was begun. Such an agent would act selectively to program cancer cells to die, as suggested by Basile et al. 11
For example, those early studies were able to show that both neuroblastoma cells 
and cells of certain leukemias can be made to mature when treated with certain naturally occurring substances, such as nerve growth factor. 12,13
An experiment using undifferentiated proerythroblasts obtained from a Friend 
erythroleukemic cell line was used to test DMSO differentiation in vitro . In the 
absence of DMSO, some gigantic cells and many undifferentiated cells were seen in culture. With the addition of DMSO to cultures, striking morphological changes were seen as cells became smaller, the nuclear–cytoplasmic ratio and nucleoli reduced, and the formation of many differentiated normoblasts occurred, suggesting that cell maturation from DMSO exposure took place, preventing these cells from turning malignant. 14
In another example, the potential by DMSO to modify leukemia cells was shown. Human acute myelogenous leukemia originates from pluripotent stem cells that transform into neoplastic cells. 15 A major phenotypic abnormality of these leukemia 
cells is their inability to differentiate to functional mature cells at the myeloblast or promyelocyte stage. Leukemic patients with this condition usually die of infection, because blast cells do not mature to their functional end cells, allowing the leukemic cells to rapidly multiply in the system. 15
More recently, it has been found that Friend erythroleukemia cells differenti -
ated by DMSO can produce hemoglobin and turn malignant cells into orthochro -
matic normoblasts.16 DMSO-treated erythroleukemic cells undergo an irreversible 
pathway to maturation and, in that process, produce globin mRNA and accumulate large amounts of hemoglobin. Hemoglobin accumulation in FLC differentiated by DMSO is needed for the erythroid maturation process. 17 What is of interest is that 
treated cells express an ability to remember (so-called memory response) the previ -
ous exposure to the DMSO treatment and to continue their differentiation even after the DMSO exposure has been discontinued. 17 This terminal maturation process of 
erythroid cells and hemoglobin synthesis remains to be further clarified. HL-60 HUMAN CELL LINE
The HL-60 human cell line was developed from peripheral blood leukocytes in a patient with acute promyelocytic leukemia, which are generally promyelocytes. 15 
This HL-60 cell line has been used as a unique in vitro  model to study various 125 DMSO in Malignancy
important aspects of human myeloid cell differentiation. When these promyelocyte 
cells underwent incubation for 5–7 days with appropriate concentrations of DMSO, they were able to differentiate into normal, peripheral blood granulocytes, despite their karyotypic abnormalities and leukemic origin. 18 These peripheral blood granu -
locytes were capable of phagocytizing bacteria and develop complement receptors.18
Treatment of human promyelocytic leukemia HL-60 cells with DMSO is known 
to cause differentiation of these cells along granulocytic/neutrophilic differentiation lineage. 19 After DMSO treatment, cultured granulocytic HL-60 cells eventually die 
via apoptosis. This process of apoptotic death induced by DMSO involves enhancing the effector proteins Bak and Bad while downregulating the apoptosis suppressor protein Bcl-2. 19 The ectopic expression of Bcl-2 protein is able to prevent apoptosis 
of granulocytes and monocyte/macrophage by a number of differentiating agents, but not when DMSO is used. Induction of apoptosis in macrophages by DMSO is of pathological significance because macrophages produce free radicals and inflamma -
tory cytokines that can damage normal cells. 20
METASTATIC LIVER DISEASE
Metastasis is the principal cause of death in cancer patients, and once it occurs, the most fundamental aspect of these cancer cells is their ability to sustain chronic growth. This growth differs from normal cell proliferation because the latter takes care to control the production and release of growth-promoting signals that ensure a homeostatic cell growth and cell number. Cancer cells have the unique ability to deregulate or block these signals and, essentially, do what they please. 21
Metastatic liver disease is a malignant tumor in the liver that has spread from 
another organ affected by cancer. Metastatic liver disease is frequently associated with primary colorectal carcinoma. More than half of the patients with metastatic liver disease will die from metastatic complications. This disease is the sixth lead -
ing cancer worldwide and the second in cancer-related deaths in the United States. 22
The reason the liver is a common site for cancer metastasis is due to its abundant 
blood supply, and since both lobes of the liver are usually involved, surgical removal is not possible unless a liver transplant is performed. Chemotherapy and a partial hepatectomy, especially in young people, is a possible treatment when only one lobe becomes cancerous. However, due to the rich blood supply of the liver, bleeding after surgery is a concern. One procedure by Salim
23 using electrocoagulation + DMSO (500 mg by mouth) 
attempted to destroy liver metastases spread from advanced colonic cancer and reduce bleeding complications. A significant 5-year survival rate was obtained with -
out major adverse effects in 78 patients with liver malignancy randomized to be treated with electrocoagulation + DMSO as compared to electrocoagulation only consisting of 77 patients. 23 The possible free radical–scavenging ability by DMSO 
was considered as a major factor in the long-term efficacy of this procedure.23
Tumor cells in culture generally grow in disarray unlike normal nontumorigenic 
cells. When these tumor cells are exposed to 1%–2% DMSO in culture, the random piling is drastically abolished within 3 days and instead, monolayers of cells in regular parallel orientation are seen to form, a pattern typical of nonmalignant fibroblasts. 24 126 Dimethyl Sulfoxide (DMSO) in Trauma and Disease
These morphological changes from malignant to nonmalignant cells in culture could 
be maintained indefinitely as long as DMSO was kept present.24
When DMSO was removed from the medium, reversal of these cells to the 
malignant state occurred after 3 days. It is possible that some of the properties that characterize DMSO, such as its stability, highly polar nature, and dielectric con-stant greater than that of water, which can enhance cell permeability, may explain, at least in part, the influence of this solvent on the hydration and solvation shells that surround the membrane components of the malignant cells and their ultimate transformation to benign cells. 24 These findings suggest that cancer is a disease of 
differentiation.25
However, the precise identification of the defect involved in the transformation 
of normal cells to malignant cells as well as the induction of cell differentiation by DMSO to normal cells is not easy to explain. Cancer cells seem to lose important control systems due to damage or loss of the genes inside the cells, a process known as mutation. Damage to the cell can be caused by a variety of factors. These factors, or carcinogens, can make the cell multiply abnormally to create cancer. DMSO COMBINED WITH ANTICANCER AGENTS
A strain of spontaneous rat lymphatic leukemia inbred rats exposed continually to low, oral concentrations of DMSO produces physiological changes in these animals that increases the therapeutic effects of the antineoplastic agent cyclophosphamide while also lowering its threshold toxicity. 26 This positive finding in a rat leukemic 
model suggests that combining DMSO with antitumor compounds in the treatment of human cancer is both feasible and attarctive. 26 It also implies that the toxicity 
seen with many of the anticancer agents may be lowered when DMSO is used as an adjuvant. This suggestion was supported by a sequential study from the same authors who 
reported that 12 anticancer agents administered to tumor-bearing rodents in the pres -
ence of 0.25% DMSO in the drinking water for over 1 year increased antineoplastic potency in 8 of these agents. 27 Several of the parenterally administered antineo -
plastic agents given with oral DMSO also inhibited the tumor growth rate in these animals. 27
Several trials of DMSO on human cancer were attempted using cyclophospha -
mide and large doses of DMSO. It was reported that when 5%–10% DMSO was given orally or intravenously to 10 patients with a brain tumor, no enhancing effects of cyclophosphamide in the cerebrospinal fluid or plasma was seen. 28 In another 
study, 15 renal carcinoma patients were given thioTEPA combined with 2% DMSO by mouth, but no beneficial effects on the tumors were noted. 29
Basal cell carcinoma is a malignant epithelial neoplasm of the skin, which 
remains the most common cutaneous malignancy in Caucasians, with as many as 2 million new cases expected to occur yearly in the United States. 30 Although basal 
cell carcinoma is not usually life threatening, it has become a medical problem of growing concern. Surgical excision remains the standard treatment for this tumor, but nonsurgical modalities also have achieved acceptable cure rates, particularly photodynamic therapy. 31127 DMSO in Malignancy
The basis of photodynamic therapy depends on a photoactivating light, a photo -
sensitizer such as 5-delta-aminolevulinic acid (5-ALA), tissue oxygen, and a target 
cell. Photosensitizers can be given systemically or locally on the skin. Topical pho -
todynamic therapy is generally well tolerated, and the usual side effects following 5-ALA include erythema and pain at the site of application. 32
Recent studies have shown that topical photodynamic therapy with 5-ALA 
and DMSO in 60 patients with basal cell carcinoma resulted in 81% of the patients becoming symptom-free 72 months after treatment with good cosmetic outcome. 33
A similar study evaluated the long-term follow-up for 19 cases of Bowen’s disease, 
a neoplastic skin disease also called squamous cell carcinoma, in which patients were given topical 5-ALA plus DMSO with a single exposure to 630 nm diode laser at differing energy dosages. At 60 months, 57% of the patients treated with 5-ALA and DMSO showed no histologic signs of squamous cell carcinoma. 34
DMSO IN EXTRAVASATION
DMSO has been used to reduce extravasation of cytotoxic agents given to cancer patients intravenously. Extravasation or leakage of these agents from the blood into the surrounding tissue is a true medical emergency that requires immediate action to prevent ulceration or tissue necrosis. The only licensed drug that is used in the United States to reverse extravasation of anthracycline, a class of drugs used in 
 cancer chemotherapy, is dexrazoxane,35 a derivative of ethylene diamine tetraacetic 
acid (EDTA), which chelates iron ions bound to anthracycline to lower the formation of superoxide radicals. 36
However, cooling and topical DMSO over the area of extravasation has been 
shown effective in reducing small anthracycline extravasations.37 The advantages of 
DMSO and cooling are the rapidity of preventing anthracycline tissue damage fol -
lowing extravasation and substantial cost-effective savings due to the high cost of a hospital maintaining dexrazoxane in its inventory. Although brief reports and anecdotal accounts have often appeared attesting the 
usefulness of DMSO in the treatment of lymphomas, glioblastomas, melanomas, and prostate and colon cancer when combined with conventional cancer therapeutic agents, these reports remain unconfirmed by strategic clinical trials. These controversial findings suggest that DMSO may have an important role in 
anticancer chemotherapy. However, further research is needed, particularly on a larger population that can be studied long term in randomized clinical trials. CANCER AND RADIATION
Radiation is a well-established carcinogen for humans, and 10% of invasive can -
cers are related to radiation.38 A study was conducted that used high linear energy 
transfer (LET) particles, which are used in radiotherapy, to irradiate a human bron-chial epithelial cell line, a stepwise neoplastic transformation of these cells occurred together with a consistent downregulation of the tumor suppressor TGFBI  gene. 39 
The irradiated cells showed fragmentation and shrinking of mitochondria, which was 128 Dimethyl Sulfoxide (DMSO) in Trauma and Disease
accompanied by a reduction in mitochondrial functions. The reduced mitochondrial 
functions were seen as lowering of cytochrome C oxidase, superoxide production, as well as succinate dehydrogenase activities when compared with nonirradiated con-trols. 39 Cytochrome C oxidase is the last enzyme in the respiratory electron transport 
chain of mitochondria and is important in the synthesis of ATP, the energy fuel of all cells. DMSO exposure of the irradiated epithelial cells was able to control the super -
oxide production and the damage caused by this free radical on the mitochondria. 39
A combination therapy using DMSO and sodium bicarbonate infusion was 
recently reported by Hoang et al.40 DMSO and sodium bicarbonate were mixed into 
a solution and infused daily for 6 months to nine patients suffering from advanced nonresectable biliary tract carcinoma, a highly fatal malignancy that has a poor response to chemotherapy. The treatment was well tolerated by all nine patients, and no adverse effects were recorded. The combined DMSO–sodium bicarbonate infu -
sion was seen to improve pain control, blood biochemical parameters, and quality of life for the patients. Notably, this treatment led to a 6-month period of progression-free survival for all nine patients in the trial, a finding the authors consider signifi -
cant of further research and application for the palliative care and extended survival of patients with this condition. 40
These findings add fuel to the conclusion that more research is needed using anti -
cancer agents mixed in DMSO solutions, a procedure that may provide more effec -
tive penetration and effectiveness in cancer suppression. Onyx EMBOLIZATION
Onyx is composed of ethylene–vinyl alcohol copolymer dissolved in DMSO and sus -
pended with micronized tantalum powder to provide contrast for visualization under fluoroscopy. Onyx liquid is used as an embolic agent of the vascular endothelium for a number of conditions including carotid-cavernous fistulas and cerebral arteriove -
nous malformations (A VMs). 41,42 Embolization of these vessels may prevent lethal 
hemorrhage if they burst or leak. Onyx has been used with radiosurgery where it was successful in 80% of patients 
treated for A VMs, showing safety and efficacy with a low rate (5%) of clinical com -
plications.43 Radiosurgery involves a single high-dose fraction of radiation directed 
stereotactically at an intracranial region of interest.44,45
The flow-directed catheters used to deliver the liquid Onyx for embolization can 
be coated with DMSO to prevent early polymerization within the catheter.46 It has 
not been investigated whether DMSO may serve a secondary role during emboliza -
tion in stabilizing A VMs besides being a vehicle for the copolymer. Transvenous occlusion of the cavernous sinus in dural arteriovenous fistula 
using Onyx embolization has been tried on patients with some success.47 Onyx with 
DMSO was injected through the superior pharyngeal branch of the ascending pha -
ryngeal artery where small transosseous feeders to the fistula arising from the supe -
rior pharyngeal branch of the ascending pharyngeal artery could be occluded by this approach especially when more conventional approaches cannot be achieved. 47
Arteriovenous fistula of the spinal dura is also amenable to Onyx endovas -
cular embolization. Although vascular malformation of the spinal cord is a rare 129 DMSO in Malignancy
clinical condition, it occurs when congestion of the spinal cord medullary venous 
plexus results in edema and neuronal ischemia.48 If not repaired, axial or radicular 
back pain, gait imbalance, weakness of the lower limbs, numbness, and bladder or bowel disturbances can result. 49
Onyx has been tried in order to embolize the abnormal communicating vessels to the 
dura and relieve the spinal cord edema that develops with a relatively high rate of suc -
cess.49 However, the neuronal ischemia is not always improved in arteriovenous fistula 
of the spinal dura using Onyx, and it is only speculative whether a high dose of intrave -
nous DMSO separate from the Onyx embolization could aid control of this condition.50
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8DMSO in Basic 
Neuroprotection
BRAIN TRAUMA OVERVIEW
The search for safe and effective agents that can reverse, arrest, or minimize the 
 consequences of traumatic brain injury has become a major research priority 
throughout the world due to the rising epidemic of brain trauma resulting from falls, 
motor vehicle accidents, sports injuries, physical assaults, and military and civilian gunshot wounds. The leading causes of traumatic brain injury in the general population are 
falls (35.2%), motor vehicle crashes (17.3%), blunt impact (i.e., being struck by or against a moving or stationary object) (16.5%), and assaults (10%). 1 These numbers 
do not include U.S. military personnel sustaining war-related injuries in Iraq and Afghanistan in the past decade. There were a total 253,330 traumatic brain injuries sustained by military personnel between 2000 and 2012; 194,561 were described as mild, 42,063 were moderate, and 6,476 were severe or penetrating wounds. 2
The Centers for Disease Control (CDC) estimated that at least 2.4 million emer -
gency department visits, hospitalizations, or deaths were related to traumatic brain injury in 2012. 3 Of these brain trauma cases, 52,000 deaths are recorded annually in 
the United States.4 Approximately 75% of these brain injuries are mild concussions 
and do not require hospitalization or aggressive treatment. Children and older adults are more likely to sustain a traumatic brain injury, usu -
ally from a fall, and it is one of the leading causes of death and disability for this segment of the population. 5 The direct and indirect economic costs of brain injury in 
the United States in 2010 were estimated at $76.5 billion annually.6
Moreover, about 3.2 million persons in the United States are living with life-long 
neurologic deficits from the consequences of head injuries.7
A workshop on head injuries, sponsored by the National Institute of Neurological 
Disorders and Stroke (NINDS), was held in May 2000 gathering some of the leading experts in the field of traumatic brain injury. 8 The group concluded that severe, closed 
head injury was a life-threatening injury that required immediate medical help and the need for the development of more effective drugs for the treatment of this trauma. It was also the consensus that none of previously tested treatments had been shown to significantly modify the outcome of severe, closed head injury. Additionally, the work -
shop members estimated that of the 50,000 patients hospitalized in the United States annually with severe, closed head injuries, 17,500 (33%) died irrespective of any treat -
ment given even when the best head injury centers were considered. 8 This was a call to 
action to urgently find more effective and safe interventions that could reduce the high fatality and permanent disabling deficits following moderate-to-severe head injuries.134 Dimethyl Sulfoxide (DMSO) in Trauma and Disease
There was also a stark warning by the workshop members that current therapies 
exhibit side effects that are potentially harmful to the patient and that can compro -
mise the outcome of trauma to the brain.8
Management of such head injuries presently relies on controlling high intracra -
nial pressure (ICP) from progressive edema buildup, increasing a low cerebral perfu -
sion, and correcting pulmonary distress. Unfortunately, no single agent is capable of 
addressing these consequences after a head injury and multiple therapy is generally required to control each problem separately. The drugs given to treat and manage the head injury complications in the past 
have been osmotic agents such as urea, glycerol, and mannitol; for control of ICP, steroids, which was given for many years until it was found to be contraindicated for head injuries due to inefficacy and increased mortality; and furosemide, a powerful diuretic also later abandoned as a treatment to lower brain pressure and swelling. If the head-injured patient survived, phenobarbital was given for the first 3 weeks after admission,
9 a protocol that has been modified in favor of outcome. Presently, barbitu -
rate coma is used temporarily, until the patient regains consciousness after which the barbiturate dose is reduced and withdrawn. The basis for using induced barbiturate coma is to reduce cerebral blood flow and metabolic demand in a patient who has sustained severe trauma to the brain. However, controversy has risen as to the benefits of barbiturate coma since it does 
not appear to prevent brain damage, possibly because the reduction of high ICP is not sustained. 10
A Cochrane analysis of randomized controlled trials of using barbiturate therapy 
and barbiturate coma showed no evidence that barbiturate therapy improved out -
come in patients with acute severe head injury.11 This analysis concluded that barbi -
turate therapy results in a fall in blood pressure in 25% of patients treated and this drug’s promotion of a hypotensive response will impair lowering ICP and improving cerebral perfusion pressure (CPP). 11 When trying to monitor the level of conscious -
ness in patients with head injury, barbiturates are impractical because they interfere with this clinical exam. Although initially effective in the control of elevated ICP, the main problems 
with the hypertonic diuretics (urea, mannitol, and furosemide) were their side effects when used repeatedly and also their tendency to rebound after multiple doses and create higher ICP than that on admission. 12,13
Hypothermia was suggested as a treatment for head injury. Although hypother -
mia can reduce ICP, the analysis of high-quality studies using therapeutic hypother -
mia showed an association with cerebrovascular disturbances, including a lowering of cerebral perfusion and an increase in morbidity on rewarming and possibly a risk of pneumonia in adult patients. 14
It was said that although these drugs saved a few from head injury death, they 
could not be correlated with the recovery outcome.9 Osmotic agents such as magne -
sium sulfate to lower ICP have had a time-honored place in neurosurgical practice 
and with the introduction, by Javid15 in 1958, of intravenous urea to treat brain swell -
ing, a new era of drugs with similar action leading to the search for better osmotics.
Dimethyl_Sulfoxide_(DMSO)_in_Trauma_and_Disease_[2015].txt
Summarize the following text section accurately and concisely. Maintain specific details and numerical information, and use anti-repetition mechanisms:

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A genetic change in a bacterium.
What is the first step in the emergence of antibiotic-resistant bacteria?
The transfer of immunity from a resistant bacterium to its offspring, ensuring the survival of subsequent generations of microbes.
What is the second way bacteria can become resistant to antibiotics?
To make otherwise ineffective antibiotics effective against multidrug-resistant bacteria.
What is the objective of searching for antimicrobial adjuvants?
The abundant blood supply in the liver, which allows cancer cells to multiply and spread.
What is the main reason for the increased risk of cancer in the liver?
There is no single effective treatment, and management relies on controlling increased intracranial pressure, increasing cerebral perfusion, and correcting pulmonary distress.
What is the current treatment for traumatic brain injury?