Sunday, October 20, 2019

You can't fool me, Nanotech

That is not an Atomic-Force-Microscopy scan measuring the thickness of a fragment of graphene film.


That is the Creature from the Black Lagoon.

Saturday, October 19, 2019

Twitch of the Death Nerve

This post was earlier cross-posted at Leonid Schneider's site, hence the unfrivolous tone. The version there is improved by Leonid's editing, background details and frame-story.


This rat is making bad decisions in a gambling task; it needs to think about its poor choices and where it went wrong. Perhaps it is distracted by algal neurotoxins or by cisplatin neuropathy, or by the removal of its teeth, or by 'visceral pain' (i.e. traumatic anal rape colo-rectal distention). Fortunately there are researchers with vagus-nerve stimulation or opto-genetic technology to end the distraction by restoring the theta-wave entrainment of its amygdala and anterior cingulate cortex! (this technology also lends itself to potential applications in grimdark cyberpunk novels).

Let me explain.

Elisabeth Bik recently blogged about the different genres of visual evidence that journal editors might expect to find enlivening a manuscript submitted to them. The "Domain of Images" [Elkins 1999] is more than microscopy and Western Blots; the Figures might feature FACS plots and histograms, and spectra in all their energy ranges, and Fourier-transformed IR spectroscopy, and X-ray diffraction patterns. These might not leave the readers any wiser if the key results are averaged values across repeated experiments, but as artisanal craft data they provide some confidence that someone did spend some time in the laboratory, But each genre also creates opportunities (and temptations!) for people to spend time in Photoshop instead, beautifying their data and airbrushing over the warts and wrinkles... or perhaps the experiments were more aspirational than actual. See also "The Subtleties a Spectrograph..." and "The mountains skipped like rams...".

And now, "Tricliceras Lobatum" has discovered an new avenue for artistic expression: nerve impulse recordings (Fig 2A, Wu et al. 2004 [3]). If that's not what happened, then perhaps someone sent a file to be printed at the same time that someone else in the lab was printing a batch of bar-codes.


But maybe 'bar-codes' are an alien concept to young people today with their 'QR codes', so for a metaphor that everyone will understand, let me draw a comparison to Renaissance siege-warfare ladders.

Now I am the first to admit my total inexperience in recording single-neuron depolarisation spikes; never in my life have I patched a clamp nor clamped a patch nor dashed a thousand kim. It was my understanding, though, that nerves and ganglia are not metronomes, nor MIDI sequencers, so they do not cycle with the clinical perfection of a Kraftwerk bassline. When there is noise in the inter-spike flat-line of the recordings, from the instrumentation or the biological whimsies of cells being cells, it should not cycle or repeat across recordings. But life is full of surprises!

Figs 2B and 2C, and C superimposed on B after a black/white reversal to see how well they cancel out

The story is centred at the University of Michigan, where Chung Owyang heads a well-appointed research group on digestion physiology, the Gastroenterology Research Unit. The emphasis there is on gut neurobiology, so no end of rats have ended their days as "preparations" - bedizened with electrodes and stomachs replumbed - providing recordings of their visceral nerve impulses and muscular contractions. His success in attracting NIH grants to UMich was recognised in 2013 with the Friedenwald Medal, "the highest honor bestowed by the American Gastroenterological Association (AGA)".
It is a tribute to Chung's vision that very soon after beginning his career at Michigan he began asking far more basic questions and established a very successful research program continuously funded by the NIH.
Sadly, his habit of "asking far more basic questions" did not extend as far as casting a cold skeptical eye over the fabricated papers set before him to sign, ripe with the aroma of cut-&-paste. Pubpeer threads document two decades of these Michigan Michigastric Michigallimaufries, with the first recorded ones filling a 2001 paper -- "Intestinal serotonin acts as a paracrine substance to mediate vagal signal transmission evoked by luminal factors in the rat" (Zhu, Wu, Owyang & Li. 2001) [1]...



...and the most recent in September: "High-fat diet–induced vagal afferent dysfunction via upregulation of 2-pore domain potassium TRESK channel" (Grabauskas et al 2019) [13].


At the risk of becoming monotonous, Figure 11, "Figure 11. Responses of the vagal pancreatic efferent nerve to microinjection of glutamate into the lateral parabrachial nucleus" from "Hypothalamic regulation of pancreatic secretion is mediated by central cholinergic pathways in the rat" (Li et al 2003) [2]. And Figure 2, "Effects of intestinal distension or an intra-arterial injection of secretin on the discharge frequency of vagal nodose neurons", from "Secretin activates vagal primary afferent neurons in the rat" (Li et al. 2005) [5].



Both have tape-loop flat-lines.

I worry about the paroxysms in "Role for NMDA receptors in visceral nociceptive transmission in the anterior cingulate cortex of viscerally hypersensitive rats" (Wu et al. 2008) [7]. Perhaps the animals contracted whooping cough.


I could continue at exhaustive length, but that would deprive the readers of the joy of strolling through the PubPeer threads themselves. Just one last recent example (for values of "recent" that include "five years ago"): this non-Euclidean monorail track comes from "Ghrelin induces leptin resistance by activation of suppressor of cytokine signaling 3 expression in male rats" (Heldsinger et al 2014) [11].


It is not clear who deserves the credit. Two names recur in the authorship list: Ying Li, and Xiaoyin Wu. Consider, for instance, "Serotonin and cholecystokinin synergistically stimulate rat vagal primary afferent neurones" (Li, Wu & Owyang 2004) [4]. Figures 1 and 5 display motoric techno beats in the baselines and repeating patterns of spikes ("Perhaps the experimental animals had hiccoughs").


Was Li claiming ownership of Fig 5 when he reprinted it as Figure 3 in his 2007 solo-authorship review paper, "Sensory signal transduction in the vagal primary afferent neurons" [A] (published in a predatory journal from Bentham)?

Turn now to "Vagal control of satiety and hormonal regulation of appetite" (Owyang & Heldsinger 2011) [9]. With neither Y. Li nor X.-Y. Wu as co-authors, Figure 3 is an exception to the earlier generalisation.


But again, this is a Review-paper issue, for Fig. 3 had earlier appeared (hiccoughs and all) as Fig 5 of "Low-affinity CCK-A receptors are coexpressed with leptin receptors in rat nodose ganglia" (Li et al, 2011) [8]. That particular output from his team must have impressed Professor Owyang as especially emblematic and worthy of a wider readership.

[8] seems to have been Li's last paper with Owyang (from 2009 onward his first affiliation was to City University of Hong Kong, and his Michigan role was downgraded to "Visiting Investigator" or "Research Fellow"). Anyway, [8] has more to offer. More synthetic entities, in Figures 1, 2 and 4...



... Then there is Figure 7. In isolation it is an elegant marker-protein-based colour map of cell specialties:

"Double fluorescence labeling of rat nodose ganglia shows expression of CCKAR and OB-Rb. A: CCKAR are stained with green fluorescent Alexa Fluor 488. B: OB-Rb are stained with red fluorescent Cy3."

But a year later, in the first post-Li paper (Heldsinger et al. (2012) [10], Figure 1 shows the same antibody-tagged cells as expressing different proteins. Meanwhile a third panel of fluorescence had been found.
H/t PubPeer contributor "Unregistered"
"Immunostaining of rat nodose ganglia (NG) neurons shows colocalization of leptin receptors (LRb), CCK-A receptors (CCKAR), and cocaine- and amphetamine-regulated transcript (CART)."

"Brycinus Lateralis" questioned whether the procedure described in the Methods of [10] could ever have produced the published results:

#4 Brycinus Lateralis
Without Leonid Schneider and Tiger to inform me, I would never have known that the critique is valid (which is why no-one is paying me to oversee a large research laboratory, nor trusting me to review submissions for AJP Gastrointestinal & Liver Physiology). The gist is that even if you have antibodies tailored to home in on (say) CCKAR and LepR, they have to be extracted from different species - say, chicken and goat. For the secondary fluorophore-conjugated antibodies (the second stage of the colour-coding) are indifferent to the target protein, and species-specific to the primary antibodies, so they will light up both distributions as red and green together.

Having slogged through that explanation (don't worry, this will not be in the test), it makes sense to look at two other double- and triple-immunohistochemistry issues. Figure 2 from Zhao et al. (2015) [12] is beautiful but meaningless:

#1 Orthogeomys Grandis


And looking at [13] again:

#1 Chapoda Fortuna
Promotion ladder is broken
One can hardly blame Ying Li for shifting his focus from Michigan to Hong Kong City University. For that NIH beneficence to the Gastroenterology Research Unit does not seem to spill over into the salaries or a promotional ladder for its staff. Departmental records show Ying Li advancing slowly from "Senior Associate Research Scientist" in 2002 to "Research Associate Professor in 2008, and from US$76,064 to $94,724. Despite her key role in the laboratory, Xiao-Yin Wu was stuck as "Research Associate" or "Research Lab Specialist" from 2002 to the present, while her salary crept slowly from US$32,240 to the still-derisory figure of $56,426. Perhaps this is standard for the precariat serfs in US academic feudalism.

Anyway, browsing through Li's Hong King output, I see the continuity from his time at Michigan... rewired rats and rat colorectal distention feature a lot, and the Anterior Cingulate cortex (the power of almost-anagrams explains the interest in ciguatera poisoning). This is where we came in.


In contrast, though, few primary data are on show in these post-Owyang papers, only the aggregate results (histograms, group-mean line plots, Fourier transforms). This may reflect the editorial preferences of different journals. At any rate it is certainly safer. I like to think that the results from Li's group are either (1) made up in their entirety so no actual rats were vivisected and tormented in the making of the movie; or (2) reported with scrupulous accuracy so that others can rely on the data and the rats did not suffer and die in vain.

A few notable exceptions to this "no raw data" rule come to mind. First, these timelines from Wang et al. (2015) [E] overlap better than one might expect.


H/t "Ophiactis Macrolepidota"
Despite some half-arsed manipulation, these protein blobs from a Western Blot - Figs 3 and 5 from Li et al (2012) [B] - remain as distinctive in their own way as Monet brushstrokes.

[B] also provides us with Figure 1(c), "Representative electromyograms of the VMR to graded-pressure CRD in VH rats treated with vehicle or glutamate, or with microelectroporation of CaMKII siRNA before glutamate injection". It seems that a thorough grounding in rewiring rats is not the only benefit Li took with him from Michigan, for the same gut-spasm electromyograms had appeared four years previously (with different labels in a different experiment) in "Anterior cingulate cortex modulates visceral pain as measured by visceromotor responses in viscerally hypersensitive rats" (Cao et al. 2008) [6]. In a third appearance in a review chapter ("Synaptic Plasticity and Synchrony in the Anterior Cingulate Cortex Circuitry", Li 2018) [F], the electromyograms reverted to their 2008 labels.


I mustn't forget the quality of 'bilocation' shown by Fig 1(a) of [B], and between Figs 1 and 4 of [6].



Two last cases of electromyogram recycling, from "Cholecystokinin enhances visceral pain-related affective memory via vagal afferent pathway in rats" (Cao et al 2012) [C] and "Vagus nerve stimulation modulates visceral pain-related affective memory" (Zhang et al. 2013) [D]:


Of course these are not fabricated figures, so each reuse could be an innocent mistake, not evidence of bad intentions. The authors can be forgiven for attentional lapses while choosing and assembling their diagrams. Perhaps they had recently incurred the removal of their molars, or been poisoned with cisplatin or algal neurotoxins, leading them to distraction and sub-optimal decision-making.

Must credit Research Affiliate TigerBB8 (current salary US$000,000.00)

Alternative Title:

Guts, guts, got no guts
And stitches don't help at all

SOURCES

[1]. "Intestinal serotonin acts as a paracrine substance to mediate vagal signal transmission evoked by luminal factors in the rat",
J. X. Zhu, X. Y. Wu, C. Owyang, Y. Li (2001)
Journal of Physiology doi: 10.1111/j.1469-7793.2001.0431k.x [PubPeer].

[2]. "Hypothalamic regulation of pancreatic secretion is mediated by central cholinergic pathways in the rat", Ying Li, Xiaoyin Wu , Jinxia Zhu , Jin Yan , Chung Owyang (2003).
Journal of Physiology doi: 10.1113/jphysiol.2003.049122 [PubPeer].

[3]. "Hypothalamus–Brain Stem Circuitry Responsible for Vagal Efferent Signaling to the Pancreas Evoked By Hypoglycemia in Rat", Xiaoyin Wu, Jun Gao, Jin Yan, Chung Owyang, Ying Li (2004).
Journal of Neurophysiology doi: 10.1152/jn.00791.2003 [PubPeer].

[4]. "Serotonin and cholecystokinin synergistically stimulate rat vagal primary afferent neurones", Y. Li, X. Y. Wu , C. Owyang (2004).
Journal of Physiology doi: 10.1113/jphysiol.2004.064816 [PubPeer].

[5]. "Secretin activates vagal primary afferent neurons in the rat: evidence from electrophysiological and immunohistochemical studies", Ying Li , Xiaoyin Wu , Harry Yao , Chung Owyang (2005).
AJP Gastrointestinal & Liver Physiology doi: 10.1152/ajpgi.00039.2005 [PubPeer].

[6]. "Anterior cingulate cortex modulates visceral pain as measured by visceromotor responses in viscerally hypersensitive rats", Zhijun Cao , Xiaoyin Wu , Shengliang Chen , Jing Fan , Rui Zhang , Chung Owyang , Ying Li (2008).
Gastroenterology (2008) doi: 10.1053/j.gastro.2007.11.057 [PubPeer].

[7]. "Role for NMDA receptors in visceral nociceptive transmission in the anterior cingulate cortex of viscerally hypersensitive rats", Xiaoyin Wu , Jun Gao , Jin Yan , Jing Fan , Chung Owyang , Ying Li (2008).
AJP Gastrointestinal & Liver Physiology doi: 10.1152/ajpgi.00452.2007 [PubPeer].

[8]. "Low-affinity CCK-A receptors are coexpressed with leptin receptors in rat nodose ganglia: implications for leptin as a regulator of short-term satiety", Ying Li , Xiaoyin Wu , Shiyi Zhou , Chung Owyang (2011).
AJP Gastrointestinal & Liver Physiology doi: 10.1152/ajpgi.00356.2010 [PubPeer].

[9]. "Vagal control of satiety and hormonal regulation of appetite", Chung Owyang, Andrea Heldsinger (2011).
Journal of Neurogastroenterology & Motility doi: 10.5056/jnm.2011.17.4.338 [PubPeer].

[10]. "Cocaine- and amphetamine-regulated transcript is the neurotransmitter regulating the action of cholecystokinin and leptin on short-term satiety in rats", Andrea Heldsinger , Yuanxu Lu , Shi-Yi Zhou , Xiaoyin Wu , Gintautas Grabauskas , Il Song , Chung Owyang (2012)
AJP Gastrointestinal & Liver Physiology doi: 10.1152/ajpgi.00231.2012 [PubPeer].

[11]. "Ghrelin induces leptin resistance by activation of suppressor of cytokine signaling 3 expression in male rats: implications in satiety regulation", Andrea Heldsinger , Gintautas Grabauskas , Xiaoyin Wu , ShiYi Zhou , Yuanxu Lu , Il Song , Chung Owyang (2014).
Endocrinology doi: 10.1210/en.2013-2095 [PubPeer].

[12]. "Upregulation of bile acid receptor TGR5 and nNOS in gastric myenteric plexus is responsible for delayed gastric emptying after chronic high-fat feeding in rats", Hui Zhou , Shiyi Zhou , Jun Gao , Guanpo Zhang , Yuanxu Lu , Chung Owyang (2015).
AJP Gastrointestinal & Liver Physiology doi: 10.1152/ajpgi.00380.2014 [PubPeer].

[13]. "High-fat diet–induced vagal afferent dysfunction via upregulation of 2-pore domain potassium TRESK channel", Gintautas Grabauskas , Xiaoyin Wu , ShiYi Zhou , JiYao Li , Jun Gao , Chung Owyang (2019).
JCI Insight doi: 10.1172/jci.insight.130402 [PubPeer].
* * * * * * * * * * * * * * * *
[A] "Sensory signal transduction in the vagal primary afferent neurons", Ying Li (2007).
Current Medicinal Chemistry doi: 10.2174/092986707782023334 [PubPeer].

[B]. "Phosphorylated CaMKII post-synaptic binding to NR2B subunits in the anterior cingulate cortex mediates visceral pain in visceral hypersensitive rats", Ying Li , Xu Zhang , Haiyan Liu , Zhijun Cao , Shengliang Chen , Bing Cao , Jin Liu (2012).
Journal of Neurochemistry doi: 10.1111/j.1471-4159.2012.07717.x [PubPeer].

[C]. "Cholecystokinin enhances visceral pain-related affective memory via vagal afferent pathway in rats", Bing Cao, Xu Zhang, Ni Yan, Shengliang Chen, Ying Li (2012).
Molecular Brain doi: 10.1186/1756-6606-5-19 [PubPeer].

[D]. "Vagus nerve stimulation modulates visceral pain-related affective memory", Xu Zhang , Bing Cao , Ni Yan , Jin Liu , Jun Wang , Vivian Oi Vian Tung , Ying Li (2013).
Behavioural Brain Research doi: 10.1016/j.bbr.2012.08.027 [PubPeer].

[E] "Facilitation of synaptic transmission in the anterior cingulate cortex in viscerally hypersensitive rats", Jun Wang , X. Zhang , Bing Cao , Jin Liu , Ying Li (2015).
Cerebral Cortex doi: 10.1093/cercor/bht273 [PubPeer].

[F]. "Synaptic Plasticity and Synchrony in the Anterior Cingulate Cortex Circuitry: A Neural Network Approach to Causality of Chronic Visceral Pain and Associated Cognitive Deficits", Ying Li (2018).
Advances in Neurobiology doi: 10.1007/978-3-319-94593-4_8 [PubPeer].

Tuesday, October 15, 2019

A turnip for the books

London police arrest man dressed as broccoli


Reuters reports, without offering any context, the fact that a man dressed as a giant broccoli was arrested yesterday in London. While being detained, he yells "give peas a chance."

Wherever a corporation is destroying the environment, Broccoli Man is there. Where there's muck there's brassica.

Friday, October 4, 2019

F-I-R-E-I-N-C-A-I-R-O

This post was earlier cross-posted at Leonid Schneider's site, hence the unfrivolous tone. The version there is improved by Leonid's editing, background details and frame-story. It is a sequel (or possibly prequel) to an earlier post.


The "Curate's Egg" joke has earned itself a special place in the history of integrity in scholarship, due to the outrage and calls for retraction, after the best-known version in a 1895 Punch turned out to have been plagiarised from a slightly earlier cartoon in a rival magazine. Not really true about the 'outrage', though please appreciate this Twiddle account, devoted to Victorian humour (the Golden Age of Dad Jokes). Here the yolk is the naive suggestion that in a homogeneous unitary product, only part might be decayed, leaving the remaining material perfectly palatable and reliable.



It may seem a stretch to get from there to Professor Gaetano Cairo's participation in PubPeer comment threads, where he has defended a half-dozen papers against criticism and hostile questioning. We should all admire his involvement, by the way, and hope that it inspires other researchers to do the same. But it is not really such a jump, for Cairo's courteous responses capture some of the curate's polyglossian positivity ('Polyglossian' is a portmanteau word I invented just now by combining 'Pangloss' with 'Pollyanna'). Their general theme is that one or two Figure components might have been taken by error from another context, or slightly fabricated by persons unknown, but the other results are above suspicion so the main conclusions of each paper remain unaltered.

For instance, "Loss of the von Hippel Lindau tumor suppressor disrupts iron homeostasis in renal carcinoma cells" (Alberghini et al. 2005) [5]. Here the objects of apologia are two loading controls (congratulations to those of you who had "Loading Control Library" in your Bingo cards!).



Murmuring 'Zoom... Enhance' like Deckard in Bladerunner, we tweak the contrast and find that the former control is an exercise in the artistic medium of decoupage.

Thank you for this remark. I would like to respectfully remark that the spot circled in red is invisible and the actin band in the yellow box would appear neatly cut if duplicated as suggested, whereas it appears rounded. The 18S are similar but also not so different from those in the adjacent lanes. Anyway, almost fifteen years after publication, the original films are no longer retrievable and thus it’s unfortunately impossible for us to evaluate them. We would like to remark that the potential image duplication, though inappropriate, does not affect the main results and conclusions of this work. In fact, the figures report a complementary experiment showing the well-known opposite regulation of IRPs and ferritins by iron and iron chelators.
Example (2) comes from "Doxorubicin paradoxically protects cardiomyocytes against iron-mediated toxicity" (Corna et al. 2004) [4]. To the casual eye, two or even three of the four loading-control lanes in Fig 3A are clones, with a few percentage variation in width, as can happen with identical triplets.


Thank you for this remark. The bands are very similar but also not so different from those in the adjacent lanes. It is actually hard for us to agree or disagree by inspection that the rRNA images were duplicated, as commented by Xanthorrhoea Gracilis. The paper is 15 years old and therefore, unfortunately, it’s impossible for us to retrieve and evaluate the original images of the ethidium bromide-stained rRNA bands used for this figure. In case of inadvertent error, we sincerely apologize, but we would like to remark that the hypothetical use of the same image for two different lanes, though inappropriate, does not affect the results and conclusions of this work.
The mystery was deepened by the emergence of the same rRNA band in Fig 3B of "Reactive oxygen species-independent apoptosis in doxorubicin-treated H9c2 cardiomyocytes" (Bernuzzi et al. 2009) [11] -- vertically compressed, and controlling across a quite different range of conditions.


It was accompanied there by another rRNA band in Fig 4A, in which the two right-hand lanes are novel (but identical) but the two left-hand lanes are ones we've already seen.

If arguendo we accept that the loading controls were assembled by cloning lanes within bands taken from elsewhere, then two corollaries follow: First, the lanes were not controlled against unintended fluctuations in the concentrations and quantities of lysate from each experimental condition. Second, whoever in the research team made up evidence to secure the desired outcome, what are the odds that they stopped after just one clue?

Crucially, the mystery manipulators may not have confined themselves to control bands. Go back to our first Instance of a problematic loading band, from Fig 4A of [5], which controlled across smeary clouds of Ft L protein (below, left). We have just met the same cloud-bank in [11], though flipped through 180°, recut in the manner of a purloined gemstone from six carats lanes down to four, and relabelled as 'HO-1'.


Other examples of repurposed bands can be found in this body of work... enough to start one wondering whether there was a single lab member in charge of pulling random blots out of the image archive when assembling a Figure, or did they share the task around on a roster? This next control appears twice in "Adenosine A(2)A receptor but not HIF-1 mediates Tyrosine hydroxylase induction in hypoxic PC12 cells" (Gammella, Cairo & Tacchini 2010) [12], as TFIID and as β-actin; in the latter manifestation it could easily be an X-ray of my index finger.


Another phalanx-bone had previously come to light in "Role of HIF-1 and NF-kappaB transcription factors in the modulation of transferrin receptor by inflammatory and anti-inflammatory signals" (Tacchini et al. 2008) [9].


[9] leads us to [8] and [13], "Adenosine A2a receptor-mediated, normoxic induction of HIF-1 through PKC and PI-3K-dependent pathways in macrophages" (De Ponti et al. 2007) and "Role of hypoxia-inducible factors in the dexrazoxane-mediated protection of cardiomyocytes from doxorubicin-induced toxicity" (Spagnuolo et al. 2011) -- the three papers being tightly intertwined despite their diverse topics. At left, [9] and [8] share a loading control which resembles a row of burning canoes.



In [9] the canoes are matched to a scene of full-rigged ships-of-the-line emerging through cannon smoke, in a little-known naval engagement from the Napoleonic Wars (above, right). In [13] the ships change their lane labels and lose one of their number, but acquire a new TFIID chaperone.

In less nautical imagery, two HIF-1 bands from Fig 1D of [8] and 1(a) of [13] are surprisingly similar after a horizontal flip:


Seriously, how does all this happen? We must consider the possibility that Prof. Cairo had only limited oversight of what was going on in the laboratory at the time of these papers.

Anyway, I haven't finished with [8]. Its Figures 6A and 7A feature a pair of Forensic Dental X-ray images, shared with Fig 5B of "Up regulation of IL-6 by ischemic preconditioning in normal and fatty rat livers" (Tacchini et al. 2006) [6]. One of those dental records is hard-working: it leads us back to 2010, with 3D of [12] (the neighbourhood of the phalangeal X-ray).


[6] rewards further attention. Within its Fig 5A, two "Oct-1" loading controls from Fig 2A are easily mistaken for a row of cat emojis, and cancel out surprisingly well when superimposed.


Their resemblance inspired 'Maianthemum Lichiangense' to draw four papers together into one diagram: Figs 1B from [8], 5A from [6], 2a from [9] and 3 from "Adenosine‐dependent activation of hypoxia‐inducible factor‐1 induces late preconditioning in liver cells" (Alchera et al. 2008) [10].


As the end of this post approaches, I belatedly realise that I never set out its purpose -- of providing some context for an earlier post. That one looked at the oeuvre of Prof. Alfonsina Desiderio, also of Milan University. This is either a sequel or a prequel. As Leonid noted in his Introduction to that post, there are historical links between the two research groups. It is not for me to wonder whether they speak of a broader research-culture malaise within Milan University.

There are also pictorial connections. For instance, this one, involving Figs 3(a) of "IRP1-independent alterations of cardiac iron metabolism in doxorubicin-treated mice" (Corna et al. 2006) [7] and 2C in a 2004 paper from Desiderio's group:


Now Prof Cairo did not find resemblance to be convincing. His frequent co-author Tacchini had collaborated with Desiderio in the latter paper, but not with his own team in the former, so the authorship lists of the two citations did not overlap:
Thank you for this remark. In reply to comments to a few other old papers I did not argue because the original data were no longer available for evaluation. However, in this case the comment of Pandanus Comatus about 18S RNA seems inappropriate because it regards two independent papers with no authors in common. I cannot see how the original image could have been duplicated.
It may be that a second image, shared between the respective Figs 2B and 3(a), is more convincing.


Finally, 'Macaranga Lowii' put together another sprawling diagram calling attention to similarities among a series of blots. The implication is that an original 10-lane band of S28 ribosomal RNA had been cut to various widths depending on the number of conditions retrospectively needing a loading control in each experiment.


Three of the five papers are currently relevant, though old enough to interest only historians... At the bottom of the chart is 'HIF-1-mediated activation of transferrin receptor gene transcription by iron chelation' (Bianchi, Tacchini & Cairo, 1999) [1]. At left, "Transferrin receptor induction by hypoxia" (Tacchini et al. 1999) [2]. At lower left, "Oxidative stress-mediated down-regulation of rat hydroxyacid oxidase 1" (Recalcati et al. 2003) [3]. There is some authorship overlap with the 2001 and 2002 papers from Desiderio's early work.


Conceding the possibility that a single band might have used more than once by mistake for loading-control purposes, Professor Cairo has pointed out that the lack of distinctiveness in the shapes of these Magritte-bilboquet blobs makes it difficult to be sure. Consulting the original data is impossible, given the largely antiquarian nature of these papers.
It is actually possible that we inadvertently duplicated the rRNA images, as commented by Streptomyces Avellaneus, though all these rRNA bands look alike. The J Biol Chem paper is 20 years old and the experiments were performed 1-2 years earlier, before the digital images era. Therefore, unfortunately, it’s impossible for us to retrieve and evaluate the original images of the ethidium bromide-stained rRNA bands used for this figure.
I agree that some of the proposed identities in the chart may be false positives, mistaken identities, But my general conclusions remain unaltered.

Sources:
[1]. "HIF-1-mediated activation of transferrin receptor gene transcription by iron chelation".
L. Bianchi, L. Tacchini, G. Cairo, Nucleic Acids Research (1999) doi: 10.1093/nar/27.21.4223 [PubPeer]

[2]. "Transferrin receptor induction by hypoxia. HIF-1-mediated transcriptional activation and cell-specific post-transcriptional regulation".
Lorenza Tacchini, Laura Bianchi, Aldo Bernelli-Zazzera, Gaetano Cairo, Journal of biological chemistry (1999) 10.1074/jbc.274.34.24142 [PubPeer]

[3]. "Oxidative stress-mediated down-regulation of rat hydroxyacid oxidase 1, a liver-specific peroxisomal enzyme"
Stefania Recalcati, Lorenza Tacchini, Alessandra Alberghini, Dario Conte, Gaetano Cairo, Hepatology (2003) doi: 10.1053/jhep.2003.50417 [PubPeer]

[4]. "Doxorubicin paradoxically protects cardiomyocytes against iron-mediated toxicity: role of reactive oxygen species and ferritin".
Gianfranca Corna, Paolo Santambrogio, Giorgio Minotti, Gaetano Cairo, Journal of Biological Chemistry (2004) doi: 10.1074/jbc.m310106200 [PubPeer]

[5]. "Loss of the von Hippel Lindau tumor suppressor disrupts iron homeostasis in renal carcinoma cells".
Alessandra Alberghini, Stefania Recalcati, Lorenza Tacchini, Paolo Santambrogio, Alessandro Campanella, Gaetano Cairo, Journal of Biological Chemistry (2005) doi: 10.1074/jbc.m500971200 [PubPeer].

[6]. "Up regulation of IL-6 by ischemic preconditioning in normal and fatty rat livers: association with reduction of oxidative stress"
Lorenza Tacchini, Lorenza Tacchini, Gaetano Cairo, Lorenza Tacchini, Gaetano Cairo, Cristina De Ponti, Marta Massip, Joan Rosellò-Catafau, Carmen Peralta, Free Radical Research (2006) doi: 10.1080/10715760600885432 [PubPeer]

[7]. "IRP1-independent alterations of cardiac iron metabolism in doxorubicin-treated mice"
Gianfranca Corna, Bruno Galy, Matthias W. Hentze, Gaetano Cairo, Journal of Molecular Medicine (2006) doi: 10.1007/s00109-006-0068-y [PubPeer]

[8]. "Adenosine A2a receptor-mediated, normoxic induction of HIF-1 through PKC and PI-3K-dependent pathways in macrophages".
Cristina De Ponti, Rita Carini, Elisa Alchera, Maria Paola Nitti, Massimo Locati, Emanuele Albano, Gaetano Cairo, Lorenza Tacchini, Journal of Leukocyte Biology (2007) doi: 10.1189/jlb.0107060 [PubPeer]

[9]. "Role of HIF-1 and NF-kappaB transcription factors in the modulation of transferrin receptor by inflammatory and anti-inflammatory signals".
Lorenza Tacchini, Elena Gammella, Cristina De Ponti, Stefania Recalcati, Gaetano Cairo, Journal of Biological Chemistry (2008) doi: 10.1074/jbc.m800365200 [PubPeer]

[10]. "Adenosine‐dependent activation of hypoxia‐inducible factor‐1 induces late preconditioning in liver cells".
Elisa Alchera, Lorenza Tacchini, Chiara Imarisio, Caterina Dal Ponte, Cristina De Ponti, Elena Gammella, Gaetano Cairo, Emanuele Albano, Rita Carini, Hepatology (2008) doi: 10.1002/hep.22249 [PubPeer]

[11]. "Reactive oxygen species-independent apoptosis in doxorubicin-treated H9c2 cardiomyocytes: role for heme oxygenase-1 down-modulation".
Francesca Bernuzzi, Stefania Recalcati, Alessandra Alberghini, Gaetano Cairo, Chemico-Biological Interactions (2009) doi: 10.1016/j.cbi.2008.09.012 [PubPeer]

[12]. "Adenosine A(2)A receptor but not HIF-1 mediates Tyrosine hydroxylase induction in hypoxic PC12 cells".
Elena Gammella, Gaetano Cairo, Lorenza Tacchini, Journal of Neuroscience Research (2010) doi: 10.1002/jnr.22366 [PubPeer]

[13]. "Role of hypoxia-inducible factors in the dexrazoxane-mediated protection of cardiomyocytes from doxorubicin-induced toxicity"
R D Spagnuolo, S Recalcati, L Tacchini, G Cairo, British Journal of Pharmacology (2011) doi: 10.1111/j.1476-5381.2011.01208.x [PubPeer]