Pensées - entry 00006

So... here. As promised in Pensées from Moi.

Original publishing date: April 13, 2010.

Original Title: Homeopathy and mouse model

Continuing to commemorate the World Homeopathy Awareness Week (WHAW)...

I came across this paper (Homeopathy, 2009, 98:267-79), which explores the utility of mouse models in homeopathy research, looked at the putative efficacy of homeopathic drugs against radiation-, cytotoxicity-, genotoxicity- and oxidative stress induced damage. I was particularly curious about this paper because of my long association with mouse models in various infectious diseases.

Note: I wish I could post parts of the figures here. But such use is restricted by the publisher. If anyone is interested, I can provide a copy of the PDF for personal academic use.

Right in the introduction, the author proposed to use mice to test the presumed efficacy of homeopathic drugs, particularly those diluted beyond the inverse of Avogadro's number (~6 x 10²³). Host damage was induced in the mice at the tissue, cellular or DNA level by treating them with toxic chemicals, carcinogens or mutagens. Homeopathic remedies (HRs) were administered, and the effects were assessed by cytology, histopathology and biochemical means.

The HRs were chosen based on their purported effects on certain disease conditions following the similarity principle, as well as organ specific action, and was given to the test group of experimentally damaged mice. The control groups consisted of similar experimental mice given (a) only 'succussed alcohol' (diluent for HRs), and (b) 'wrong' HR, which is not specific for the organ.

Experimental outcomes were dichotomized as 'positive' and 'negative', and expected 'positive' results were defined as any statistically significant difference observed in the test groups compared to controls. The author took pains to indicate that not all mice in the test group may respond equally to HR, or show amelioration of their symptoms.

My curiosity now piqued, I delved into the paper. A large array of techniques was cited in the Methods section: various cytogenetic assays, including micronucleus induction assay (which enumerates the level of chromosomal damage), sperm-head morphology assay (for spermatotoxicity), mitotic index assay (which identifies effect of substances on cell proliferation), the Comet assay (which evaluates DNA damage/repair), and flow-cytometric analysis of cell cycle; Western and RT-PCR based analysis of the expression of various signaling molecules, and cell-cycle and oncogene-associated proteins, followed by immunohistochemistry; assay for various enzymatic biomarkers of hepatic function (such as phosphatases and transaminases) and toxicity; histopathology and EM.

The methods section, full of verbose explanation of techniques, was surprisingly short of description of actual reagents used. The results section appeared to be a summary of various papers published by the same group (with references indicated in lieu of actual evidence), which is rather odd for a paper written in a format as if presenting primary data. Anyway, undeterred I plodded along, trying to decipher the undefined abbreviations used.

The credibility of the data in Table 1 and 2 (the only places - apart from a couple of figures - where original data were reported) comes under a heavy cloud based on several crucial points. Based on author's descprition, the data were presumably collected in a pooled manner from 5-6 mice, but the experimental results represent mean of triplicates (hence, N = 3, a very small sample size). The Tables 1 and 2 indicated that they contained hitherto unpublished data from studies that used Dimethylbenzanthracene (DMBA), a potent carcinogen, and Croton oil, a skin irritant and exfoliant, to create skin papillomas in mice.

Contrary to the experimental description, only untreated test group, HR (Kalium Arsenum 30C)-treated test group, and control group 'a' results (but not control group 'b') were shown along with those from normal mice. Also, the displayed values didn't include all the parameters mentioned in the methods section. Not only that, there was no mention of the affected cell type in which the assays were done. For example, Croton oil causes diarrhea when ingested orally, and it was mentioned that the HRs and the control substances were delivered by oral gavage. How the orally consumed HR was targeted to and associated with specific organ/tissue locations - particularly in a skin papilloma model - were not indicated.

Given the small sample size, the observed differences between control and test groups seem rather small and biologically irrelevant (though statistical significance was claimed in favor of the HR treatment), especially since the description of the tissue/cells assayed was left obscure; I am tempted to suspect a massive amount of confirmation bias in scoring the observations. The study doesn't have enough power to dispel that notion. A summary of Tables 1 and 2 (since the original Tables are copyrighted by the publisher, Elsevier) shows this (showing only means for brevity's sake):

Group	Caspase3 activity	Micro- nuclei %	Mitotic index %	SOD Enzyme	Catalase	Lipid Peroxidation	Glutathione
Carcinogen	14.4	1.1	7.4	9.8	26.1	42	23.3
Carcinogen + Alcohol	12.8	1.2	7	8.3	23.7	45	22.3
Carcinogen + HR	16.7	0.7	3.8	14	29.1	40	27.9

In addition, Western Blots and RT-PCR products for various signalling, cytokine and oncogene-associated products shown in Figure 2 are not very convincing as to the difference between groups in absence of information of amounts of protein or DNA loaded to run the gels. Despite the author's claims, the histopathology and immunohistochemistry shown in Figure 3 did not appear to show any discernible difference between the control group 'a' and the HR-treated test group.

The rest of the result section also talked about studies in HR-induced immunomodulation, without supplying any evidence in corroboration of their claims.

A similar trend of tall claims bereft of evidence was observed in the discussion section. In trying to explain the 'observations', the author fell back on that tired, old, implausible canard of 'water memory' a.k.a. 'imprint':

... The leading current proposals for the mode of action of such 'ultra molecular' dilutions (Note: it means dilutions beyond Avogadro's limit) is that water is capable of storing information relating to substances with which it has been in contact and subsequently can transmit this information to pre-sensitized biological system. The process is believed to be mediated by structural modification of water, analogous to storage of information by magnetic media...

Next, the author advanced a hypothesis that homeopathic dilutions worked by modulating gene expression, including those associated with cell signaling and cytokines. After acknowledging that the prospect of ligand-receptor interaction becomes questionable at homeopathic dilutions, the author proposed another hypothesis:

... One possibility is that homeopathic medicines, during the 'dynamization/potentization' process, interact with nanoparticles that eventually may have some role in tagging onto some proteins, that eventually act as ligand...

... once again, without providing much by way of corroborative evidence. Perhaps the author needs to be reminded of those golden words by Carl Sagan, "Extraordinary claims require extraordinary evidence."

One major aspect of this paper that stood out immediately was the sheer amount of science-y sounding verbiage used; it was almost as if lengthy explanations of the use of techniques and terminology liberally borrowed by the fields of cytogenetics could make up for the absence of real, hard data to corroborate the claims of efficacy made in the paper.

The second aspect that struck me was the rather awfully poor quality of references (including ones that have been repeatedly cited in the paper), which included conference proceedings, unpublished PhD theses and about 35 papers (out of 77) from the author's own group (a move that seemed almost academically incestuous) published in obscure journals, including complementary medicine journals of dubious provenance, a few other papers from collaborating groups published in Homeopathy and similar journals, with a smattering of legitimate papers thrown in here and there in support of the verbiage.

Conclusion? Yawn. Moving along...