MiR-532-5p acts as a tumor suppressor and inhibits glioma cell proliferation by targeting CSF1
Authors: D Liu
J Liu
X-D Wang
Y-P Wang
A-M Bian
X-B Hui
D-Z Fang
Journal: European review for medical and pharmacological sciences (2019)
Discipline: PHARMACOLOGY & TOXICOLOGY
Keywords: miR-532-5p; glioma development; molecular functions; tumor; carcinogenesis
Affiliations: Department of Neurosurgery, The Affiliated Huaian No. 1 People's Hospital of Nanjing Medical University, Huaian, China. .

Abstract:  Objective: Recent studies have discovered a class of micro-RNAs (miRNAs), which are dysregulated in various tumors and associated with carcinogenesis. In our research, we aim to uncover the molecular functions of miR-532-5p in glioma development. Patients and methods: Real Time-quantitative Polymerase Chain Reaction (RT-qPCR) was performed to detect miR-532-5p expression in 48 glioma samples and 4 glioma cell lines. The Pearson's Chi-square test was used to determine the association of miR-532-5p expression with several clinicopathological indexes in glioma patients. Besides, cell proliferation assay, colony formation assay, and Ethynyl deoxyuridine (EdU) incorporation assay were performed to explore in vitro effects of miR-532-5p on glioma cells. Furthermore, the interaction between miR-532-5p and CSF1 in glioma was studied by performing Western blot assay and Dual-Luciferase Reporter Gene Assay. Results: Downregulated miR-532-5p expression was observed in glioma tissues compared with adjacent normal samples. MiR-532-5p expression was associated with the KPS score and tumor grading in glioma patients. Moreover, cell proliferation of glioma was inhibited after overexpression of miR-532-5p in vitro. Furthermore, CSF1 was a target of miR-532-5p in glioma. After overexpression of miR-532-5p, CSF1 was downregulated at mRNA and protein levels in vitro Besides, the expression of CSF1 in glioma tissues was negatively related to that of miR-532-5p. Conclusions: Malignant phenotypes of glioma cells were remarkably suppressed through the overexpression of miR-532-5p. MiR-532-5p/CSF1 axis was identified as a new therapeutic intervention for the treatment of glioma. 

Retracted: Yes

PubPeer comments:
null, 
 # A fundamentally flawed and misleading meta-analysis **Introduction** The paper has been widely cited to argue that e-cigarettes reduce the smoking cessation rate and therefore prolong smoking. This argument has found a willing audience in anti-vaping campaigners worldwide. The study is, however, fundamentally flawed. The fatal problem with this study is the *heterogeneity*, which is another way of saying that studies with completely different designs, exposures, outcomes and study populations have all been lumped together using a technique that is only supposed to be used to combine studies that are *very similar* in design (e.g. multiple RCTs with similar exposures, study populations and outcome measures) to improve overall statistical power. So we have studies where e-cigarette users included may have only tried the product once, where they weren t trying to quit smoking at all, and where they were not included if they had already quit smoking - all introducing a range of biases that render the meta-analysis meaningless and misleading. **Major criticisms of the paper** Professor Peter Hajek criticises the study as follows: [1] u201cThis review is grossly misleading in my opinion. There are several problems with the way studies were selected and used, but the main flaw is simple, though not easy to spot. The studies that are presented as showing that vaping does not help people quit only recruited people who were currently smoking and asked them if they used e-cigarettes in the past. This means that people who used e-cigarettes and stopped smoking were excluded. The same approach would show that proven stop-smoking medications do not help or even undermine quitting. For an academic treatment of the failings of the Kalkhoran Glantz meta-analysis, please see Villanti et al. 2018 [2]. Citing this study at reference [17], Villanti et al. say: The other two published meta-analyses included heterogeneous studies from RCTs, longitudinal and cross-sectional designs in their pooled analyses [17,23]. There are three key issues related to combining results from non-randomized studies with respect to adjustment for confounding. First, effect estimates and standard errors from non-randomized studies do not correct for imbalance in the exposed and unexposed groups (as in randomized studies). Secondly, nonrandomized studies on the same topics are likely to control for different confounders in their analyses; this creates further heterogeneity in the included studies. Thirdly, adjustment for confounding in individual studiesu2014and in pooled analysesu2014may yield a more precise estimate, but does not reduce bias. The Cochrane Handbook warns: u2018meta-analyses of studies that are at risk of bias may be seriously misleading. If bias is present in each (or some) of the individual studies, meta-analysis will simply compound the errors, and produce a u2018wrongu2019 result that may be interpreted as having more credibilityu2019 **Original co-author now accepts the study s conclusions are flawed and the analysis misleading** By 2019, the original co-author Dr. Sara Kalkhoran, by now in a different university, refutes the validity of her earlier study. She and her co-authors come to a different conclusion in their 2019 study [3] In this longitudinal cohort study of U.S. adult cigarette smokers, daily but not non-daily e-cigarette use was associated with higher odds of prolonged cigarette smoking abstinence over two years, compared to no e-cigarette use. Daily use of e-cigarettes may help some smokers to stop smoking combustible cigarettes. By way of explanation for the difference, Kalkhoran, Chang Rigotti, 2019 draw similar observations to Villanti et al., 2017 about Kalkhoran Glantz, 2016, which is citation 26 in the 2019 study: In contrast to our findings, several prior observational studies examining the relationship between e-cigarette use and cigarette abstinence found that e-cigarette use was associated with a lower likelihood of successful cessation. [26] This is likely due to heterogeneity in the way that these prior studies defined current e-cigarette use. Our findings suggest that combining daily and non-daily e-cigarette users is likely to underestimate the association between e-cigarette use, and subsequent cigarette abstinence and may miss differences in smoking abstinence among subgroups of e-cigarette users. Observational studies should account for the frequency of e-cigarette use when evaluating the association between e-cigarette use and cigarette smoking abstinence. **The more reliable evidence points to the opposite conclusion** The conclusions of the study contrast with the body of evidence from credible studies that meet more rigorous standards. in 2018, Abrams et al. contrasted  *this problematic meta-analysis* with the prevailing base of sound evidence: [3] In sharp contrast to this problematic meta-analysis, studies that take into account how and why e-cigarettes were used (e.g., frequency and duration of use, type of device, use specifically for cessation) suggest that daily vaping can facilitate quit attempts and cessation.61u201364 Newer tank, mod and pod systems that are more satisfying (sweet spot) may improve outcome efficacy.109 Recent studies using large national US samples as well as the conclusions from Warner (2018) and the NASEM report15u201317,110 indicate that use of e-cigarettes is associated with smoking cessation and with a greater number of quit attempts than NRT.65,111u2013114 Warner and Mendez (2018)94 reported that in the UK,115,116 e-cigarettes increased smoking cessation by at least 8% and in the US by at least 12% based on studies done by Zhu et. al (2017) and others.46,111u2013113 The recent and more methodologically sound studies (see Villanti et. al for details)46 seriously challenge and debunk the conclusions of the meta-analysis of Kalkhoran and Glantz (2016)100 and the updated meta-analysis of Glantz and Bareham (2018).117 **Concluding remarks** Professor Robert West, the editor of the journal *Addiction* joined several others making numerous criticisms of the paper. West concluded [5]: u201cPublication of this study represents a major failure of the peer review system in this journal.u201d However, the study drew a specious but enduring conclusion for which there is no evidence and much that suggests the opposite: As currently being used, e-cigarettes are associated with significantly less quitting among smokers. At the time of writing (February 2020), this paper has been [cited 197 times in PubMed Central s count] . Yet it remains, and always was, deeply misleading and should be withdrawn. **References** [1] Peter Hajek, Expert reaction to meta-analysis looking at e-cigarette use and smoking cessation, Comment to Science Media Centre in response to the publication publicity. January 2015 [[link] ] [2] Villanti AC, Feirman SP, Niaura RS, Pearson JL, Glasser AM, Collins LK, et al. How do we determine the impact of e-cigarettes on cigarette smoking cessation or reduction? Review and recommendations for answering the research question with scientific rigor. Addiction. 2017 Oct 3; [[link] ] [3] Kalkhoran S, Chang Y, Rigotti NA. Electronic Cigarette Use and Cigarette Abstinence Over Two Years among U.S. Smokers in the Population Assessment of Tobacco and Health Study. *Nicotine Tob Res*. 2019 Jul 11; [[link] ] [4] Abrams DB, Glasser AM, Villanti AC, Pearson JL, Rose S, Niaura RS. Managing nicotine without smoke to save lives now: Evidence for harm minimization. Prev Med (Baltim). Academic Press; 2018 Jun 23; [[link] ] [5] Robert West, Expert reaction to meta-analysis looking at e-cigarette use and smoking cessation, Comment to Science Media Centre in response to the publication publicity. January 2015 [[link] ] , 
 #5 Per the quote from Prof. Hajek above, u201dThis means that people who used e-cigarettes and stopped smoking were excluded.u201d If this is accurate, I donu2019t understand why the paper should stand without a correction. Could the last author respond? ,