Neuropathic pain is believed to arise from damage to nociceptive C fibres in diabetic neuropathy (DN). We have utilised corneal confocal microscopy (CCM) to quantify the severity of small nerve fibre damage in relation to the severity of neuropathic pain and quality of life (QoL) in patients with and without painful DN. 30 controls and patients with painful (n = 78) and painless (n = 62) DN underwent assessment of large and small nerve fibre function, CCM, neuropathic symptoms  and QoL . Patients with painful compared to painless DN, had comparable neurophysiology and vibration perception, but lower corneal nerve fibre density , branch density , length , inferior whorl length  and cold sensation threshold  and higher warm sensation threshold  indicative of small fibre damage. There was a significant association between all CCM parameters and the severity of painful neuropathic symptoms, depression score and QoL. CCM identifies small nerve fibre loss, which correlates with the severity of neuropathic symptoms and reduced QoL in patients with painful diabetic neuropathy. The true prevalence of painful diabetic neuropathy (DN) may be even higher, as most patients either do not report their symptoms or attribute them to diabetes3 and there is disagreement between the patient’s and physician’s perception of pain4. Neuropathic pain usually manifests as “tingling”, “pins and needles”, deep and dull aching, burning or electrical sensation in the feet and lower limbs, with a reduced ability to perform daily activities, nocturnal exacerbation with disturbed sleep resulting in lower mood, anxiety, depression and reduced quality of life1.Pain is one of the most disturbing symptoms that accompany diabetic peripheral neuropathy and can critically affect the patient’s quality of life in about one-third of patients with diabetic neuropathy5, autonomic dysfunction6, impaired skin microcirculation7 and epineurial blood flow8, altered sodium9 and calcium channel expression10, central sensitization11 and alterations in the spinal cord and brain13.The underlying mechanisms of painful DN are complex and include distal small fibre degeneration with regeneration14 including neuropathic pain scale (NPS)15, small fibre neuropathy symptom inventory questionnaire (SFN-SIQ)16 and quality of life (QoL) through the hospital anxiety and depression scale (HADS)17 and the 36-item short form health survey (SF-36)18. Although, pain questionnaires are easy to administer, they have limited sensitivity and low reliability19. Objective tests such as nerve conduction studies, quantitative sensory testing (QST), skin punch biopsy and CCM can help to define underlying nerve damage. Two large neuropathy phenotyping studies have shown that nerve conduction studies do not differentiate painful from painless neuropathy20, although thermal thresholds were associated with the severity of painful diabetic neuropathy21. With regard to structural damage to small fibres, reduced intraepidermal nerve fibre length22 and increased regeneration and axonal swellings have been found in patients with painful DN23. Corneal confocal microscopy (CCM) is a rapid non-invasive surrogate marker for small nerve fibre damage in diabetic peripheral neuropathy24. In two small studies we and others showed greater corneal nerve fibre loss in the central cornea25 and inferior whorl26 in patients with painful compared to painless diabetic neuropathy.The diagnosis of painful DN is challenging and subjective, based on establishing neurologic deficits and identifying the type and severity of pain through validated questionnairesThis study aimed to establish if CCM can differentiate patients with painful compared to painless DN and whether the severity of corneal nerve damage was associated with the severity of neuropathic pain and quality of life.Controls were age-matched with the patients with diabetes mellitus. Blood pressure (BP) was comparable between groups. The duration of diabetes was comparable between patients with painless and painful DN. 58% of patients with PDN were on pain medications. BMI (P < 0.005) and HbA1c (P < 0.0001) were significantly higher and total cholesterol (P < 0.0001), LDL (P < 0.05) and HDL (P < 0.04) were significantly lower in patients with painful DN compared to healthy controls. Patients with painless DN had a significantly higher HbA1c (P < 0.0001) and lower total cholesterol (P < 0.0001) compared to healthy controls. Total cholesterol (P = 0.03) and triglycerides (P = 0.01) were significantly higher in painful compared to painless DN  was significantly higher in painful and painless DN compared to healthy controls and was also significantly higher in painful compared to painless DN (P = 0.05). Sural nerve conduction velocity  and amplitude  were significantly lower and VPT  was significantly higher in patients with painful and painless DN compared to healthy controls, but did not differ between patients with painful and painless DN. WT  was significantly higher in patients with painful and painless DN compared to healthy controls and differed between painful and painless DN (P = 0.004). CT (21.35 ± 0.99 vs 26.08 ± 0.5 vs 27.67 ± 0.36) was lower in painful DN compared to painless DN (P < 0.00001) and controls (P < 0.0001) , CNBD , CNFL  and IWL  , CNBD  and IWL . Also, there were significant positive correlations between the average SF-36 score with CNFD , CNBD , CNFL  and IWL  and significant negative correlations between the severity of depression measured using the HADS questionnaire with CNFD , CNBD , CNFL  and IWL . Significant inverse correlations were found between the severity of neuropathic symptoms assessed using the SFN-SIQ questionnaire with CNFD , CNBD , CNFL  and IWL . With the pre-R-ODS questionnaire, there were significant positive correlations between the dimensions of carry, move, handle the object with CNFD  and IWL . Walking and movement correlated significantly with CNFD , CNBD , CNFL  and IWL  and between self-care with CNFD  and between eating and CNFD , CNBD  and CNFL . There was a correlation between household activities with CNBD  and CNFL  and between IWL and daily tasks  and the ability to change and hold body position .The most common descriptors of neuropathic pain among patients with painful DN assessed with the NPS questionnaire were: ~95% suffered from unpleasantness and intensity, ~89% reported deep pain, ~70% reported itchiness and ~65% cold pain. Almost every individual experienced more than one type of pain. Based on the SFN-SIQ questionnaire, the most common symptoms among patients with painful DN were dry mouth (~72%), dizziness (~67%), diaorrhea (~67%) and sensitive skin (~63%) Fig. . 83% of Patients with painful DN had a significantly reduced QoL compared to painless DN based on the SF-36 score . The HADS depression  and anxiety score  was higher in patients with painful compared to painless DN Table . PatientQoL was assessed using the pre-R-ODS questionnaire Table , Fig. 3c27. The exact cause of neuropathic pain remains elusive28, especially as pain can be experienced when there is little or no existing damage to nerves and a patient with severe neuropathy may not have any complaints29. Previous studies have reported that damage to Aδ and C fibres is associated with neuropathic pain31. CCM can quantify small fibre pathology and stratify the severity of DN32. In the present study, we demonstrate greater small fibre dysfunction (warm and cold thresholds) and damage to corneal nerve fibres but no difference in large fibre measures such as VPT and nerve conduction studies in patients with painful compared to painless DN. This agrees with our recent study showing reduced corneal nerve parameters in patients with painful compared to painless DN and an association with the severity of pain26.Painful diabetic neuropathy has a major impact on the patient’s quality of life as a result of associated anxiety and depressionet al.7 previously reported a greater reduction in corneal nerve fibre length and intra-epidermal nerve fibre length in patients with painful compared to painless DN22. Wang et al.25 reported significant alterations in corneal nerve morphology and a direct correlation with the severity of pain25, whereas Cheng et al.23 reported axonal swelling in patients with painful compared to painless DN23.Quattrini et al.21 who reported significantly worse cold and warm detection thresholds in patients with mild and moderate to severe neuropathic pain compared to patients without neuropathic pain21. Themistocleous et al.20 reported a significant abnormality in QST parameters in patients with neuropathic pain but found no difference in nerve conduction studies, intraepidermal nerve fibre density and cold pain threshold20. The current study strengthens the evidence of greater small fibre loss and dysfunction in patients with painful compared to painless neuropathy.This study also demonstrates a significant reduction in cold sensation threshold and an increase in warm sensation threshold in patients with painful compared to painless DN, consistent with Raputova 33, allowing us to assess the effect on patients’ lives including their mood, daily activities and social life. We show a significant correlation between measures of corneal nerve damage, especially at the inferior whorl with the presence and severity of neuropathic pain, quality of life and severity of depression. However, this weak association should be interpreted cautiously and suggests that factors other than the extent of small fibre damage contribute to a worse QoL and severity of pain. Patients with painful DN also suffered from greater anxiety and depression compared to those with painless DN. Previously, Gore et al.34 also demonstrated higher scores of anxiety and depression in patients with more severe neuropathic pain34. Dizziness and dry mouth were the most common symptoms among patients with painful DN and could be attributed to concomitant use of medications to relieve neuropathic pain. The most common type of pain reported was deep pain, whilst the least common was itchy pain. Others using NPS have also reported sharp and deep pain to be common whilst itching and cold pain were infrequent35. Using the pre-R-ODS questionnaire we show a significant reduction in all dimensions of QoL except eating. The SF-36 average score indicated a significant reduction in QoL  in patients with painful compared to painless DN.Validated questionnaires are an important tool for the assessment of neuropathic pain and its effect on the patients quality of lifeTo our knowledge this is the first study to evaluate the association between the severity of small fibre neuropathy using CCM and its relationship to the severity of painful symptoms, patient mood, daily activities, social life and overall QoL. Although the VAS is a validated scale for the assessment of the severity of pain, we acknowledge it is very subjective. Another limitation of the current study is the lack of validated neuropathy specific QoL tools such as NeuroQoL and Norfolk QoL-DN that more accurately quantify QoL in diabetic neuropathy.In conclusion, we show that corneal confocal microscopy detects greater corneal nerve fibre loss in patients with painful diabetic neuropathy and this correlates with the severity of neuropathic pain, the patient’s mood and QoL. These data encourage further studies to assess the potential utility of CCM in relation to a change in the severity of neuropathic pain and quality of life in clinical intervention trials.140 patients with type 1 (n = 41) and type 2 diabetes (n = 99) and 30 age-matched healthy controls were studied. All participants underwent demographic and clinical examination, detailed assessment of peripheral neuropathy and ophthalmic examination. Participants with a history of malignancy, vitamin B12 or folate deficiency, chronic renal and/or liver impairment, connective tissue or infectious disease, neuropathy of other cause than diabetes, current diabetic foot ulcer, corneal trauma, or systemic disease of the cornea and contact lens wear were excluded from the study. A signed consent form was obtained from each participant prior to taking part in the study. This study adhered to the tenets of the Declaration of Helsinki and was approved by Greater Manchester East Research Ethics Committee.36.BMI, blood pressure, lipid profile and glycated haemoglobin (HbA1c) were measured in each participant. Neurological deficits were measured using the simplified Neuropathy disability score (NDS) (0-10) and patients were confirmed to have DN based on an NDS > 2. Further, based on the visual analogue scale (VAS), patients were divided into two groups of painful (VAS > 4) (n = 78) and painless DN (VAS ≤ 4) (n = 62)Vibration perception threshold (VPT) was measured with a Neurothesiometer , and cold and warm perception thresholds were assessed with the neurosensory analyzer  on the dorsum of the left foot. In addition, sural sensory nerve amplitude and nerve conduction velocity were assessed by a consultant neurophysiologist using nerve conduction testing machine .37. Central corneal nerves and the inferior whorl were imaged at the level of sub-basal nerve plexus. CCM provides two-dimensional images with a resolution of 10μm and a size of 384 × 384 pixels.All participants were examined using slit-lamp biomicroscopy to eliminate the presence of ocular surface infection, corneal ulceration and narrow angle glaucoma prior to CCM examination. Both eyes were examined with a laser-scanning CCM  using our previously established protocol39. All corneal parameters were quantified using manual purpose-designed software . Corneal nerve fibre density (CNFD) (no/mm2), branch density (CNBD) (no/mm2), fibre length (CNFL) (mm/mm2) and inferior whorl length (IWL) (mm/mm2) were quantified.Six images from the central cornea and four images from the inferior whorl were selected following our established protocolsQoL was assessed using the validated Short Form 36 Health Survey (SF-36) score, preliminary Rasch-built Overall Disability Scale (pre-R-ODS), Hospital Anxiety Depression Scale (HADS) and small fibre neuropathy and symptom inventory questionnaire (SFN-SIQ). 96 patients completed all questionnaires.18. Our analysis included a total score, which was an average value of all 8 dimensions.SF-36 consists of 36 questions, divided into 8 dimensions and scored in a range 0-10040. Each question was scored as 0 – not possible, 1 – possible with effort and 2 – easy to perform. However, in this study, we could not use Rasch model to interpret the pre-R-ODS questionnaire, as the number of participants who completed the questionnaire was less than the number of questions, hence not fulfilling Rasch model requirements. Therefore, the pre-R-ODS questionnaire was divided into ten different dimensions: change and hold body position, walking and movement by transport, carrying, moving, handling objects, self-care, eating, household, daily tasks, meeting other people and work, studies and hobbies. For each patient, the average score for each dimension was established and expressed as a percentage.A 146-item preliminary Rasch-built Overall Disability Scale (pre-R-ODS) was used to assess the different aspects of a patient’s QoL17. The questionnaire consisted of 14 items and was divided into 2 subscales: 7 questions for the assessment of depression and 7 questions for anxiety. Each question was scored in a range of 0-3; the higher number represented the more severe symptoms. The subscale score was summed up from each question (range 0-21) and an average depression and anxiety score was compiled.HADS was used to score depression and anxiety levels16. For each participant, the overall score for SFN-SIQ was obtained from the sum of all items.Other neuropathy symptoms including dry mouth, dizziness, dry eyes, changes in sweating, were assessed using the validated small fibre neuropathy and symptom inventory questionnaire (SFN-SIQ). SFN-SIQ consisted of 13 items. For each item, the scale range was 0-3 where 0=never, 1=sometimes, 2=often and 3=always15.Neuropathic pain scale (NPS), a scale that consists of 10 items, each on a scale ranging from 0-10 was used to assess the most common types of pain among patients with painful DN(post-hoc Bonferroni for parametric parameters and LSD for non-parametric parameters) was used. Independent-samples T-test was used to compare mean values between two groups (Mann-Whitney U test for non-parametric). Pearson correlation coefficient (Spearman’s rank correlation coefficient for non-parametric) was measured to evaluate the association between variables. Graphs were made using GraphPad Prism software .IBM SPSS statistic software  was used for the analysis. All values were calculated as mean ± standard error (SE). The significance was considered as P < 0.05. To compare values between more than two groups, One-Way Anova Supplementary Information.


In this study we used a combination of measures including regional cerebral blood flow (rCBF) and heart rate variability (HRV) to investigate brain–heart correlates of longitudinal baseline changes of chronic low back pain (cLBP) after osteopathic manipulative treatment (OMT). Thirty-two right-handed patients were randomised and divided into 4 weekly session of OMT (N = 16) or Sham (N = 16). Participants aged 42.3 ± 7.3 (M/F: 20/12) with cLBP (duration: 14.6 ± 8.0 m). At the end of the study, patients receiving OMT showed decreased baseline rCBF within several regions belonging to the pain matrix , sensory regions , middle frontal lobe and left cuneus. Conversely, rCBF was increased in right anterior insula, bilateral striatum, left posterior cingulate cortex, right prefrontal cortex, left cerebellum and right ventroposterior lateral thalamus in the OMT group as compared with Sham. OMT showed a statistically significant negative correlation between baseline High Frequency HRV changes and rCBF changes at T2 in the left posterior insula and bilateral lentiform nucleus. The same brain regions showed a positive correlation between rCBF changes and Low Frequency HRV baseline changes at T2. These findings suggest that OMT can play a significant role in regulating brain–heart interaction mechanisms. Several clinical studies explored the effects of osteopathic manipulative treatment (OMT) in different pain conditions and specifically in low back pain (LBP). OMT showed significant benefit on pain intensity2, functional disability3, health-related quality of life4, functional recovery5 and drugs consumption7. A recent systematic review showed that the use of OMT is clinically effective in pain relief  and functional status  in acute and chronic nonspecific LBP8. These data led to the updating of US Guidelines for OMT in LBP patients, establishing the clinically relevant effects of OMT9. However as suggested by the Task Force on the Low Back Pain Clinical Practice Guidelines “more research is needed to understand the mechanics of OMT and its short- and long-term effects”9.Osteopathy is a wide-spread complementary medicine that steadily increased during the last decade, especially in pain treatment19.Short-term neurobiological effects seem to show that OMT has a peripheral parasympathetic anti-inflammatory effect20.Despite these clinical studies, only few papers were published investigating the effect of OMT using MRI. Recently, using Blood Oxygen Level Dependent (BOLD) fMRI, Cerritelli and colleagues showed that 4 sessions of osteopathic treatment induce a specific and distinct effect on brain activation in areas related to interoception  as well as in the right middle frontal gyrus and left striatum21. Tamburella and colleagues showed an effect of OMT on rCBF, indicating that brain perfusion was reduced in the posterior cingulate cortex and superior parietal lobule immediately after treatment, implying a possible involvement of the central autonomic network, indicating a potential central role of autonomic nervous system (ANS). However, the authors stressed the need to investigate the effects of OMT on both the ANS and CBF. The current study is designed to meet this need by using Arterial Spin Labeling (ASL) to assess regional brain perfusion, and simultaneous measurement of cardiac and respiratory activity to derive ANS indexes such as the heart rate variability (HRV).However, there is only one study that used MRI and regional cerebral blood flow (rCBF) to investigate the post-treatment effects of OMT in asymptomatic volunteers24, it is unable to offer a quantitative measure of brain activity associated to a single condition. In this regard, BOLD is severely limited when it is not possible to alternate control and experimental conditions. This is particularly true for chronic pain studies, where it is not possible to switch on and off different pain levels at will as required by standard BOLD paradigms. In contrast, noninvasive measurement of regional CBF using ASL25, offers the opportunity to obtain a measure of a well-defined physiological variable that can quantify both baseline and task induced variation of brain activity. Importantly, CBF can be measured for each condition independently, even when they are separated by days or months with good long term reproducibility26, making ASL a technique naturally suitable to study chronic pain or aging29. Thus, examining changes in rCBF over time has been considered a relevant option to study pain processing31. In this study we use a combination of measures including rCBF and HRV to investigate brain and heart rate correlates of changes in baseline levels of chronic LBP (cLBP) after OMT.Although fMRI based on the BOLD contrast can detect subtle signal variations due to graded stimulation with high sensitivity32.A recent meta-analysis of fMRI data reported that chronic pain patients showed a major role of the following structures related to the pain matrix: bilateral precentral (BA6&BA44) and postcentral gyri (BA40), bilateral insula, left S2, left dorsal ACC (BA32), left cingulate gyrus, right medial frontal gyrus (MFG), left anterior cerebellum, bilateral basal ganglia and bilateral thalamus32.According to Tanasescu and colleagues, the most frequently reported brain areas involved in chronic pain were the right posterior insula (BA13) , the left BA13 and the bilateral inferior parietal lobule  in 37–50% of all studies. About a third of the study reported bilateral putamen as well33. Indeed, Beissner et al. synthetized, in a seminal paper, the features of the so called central autonomic network (CAN), which controls the sympathetic and parasympathetic divisions that have largely divergent specific regulatory networks, differentially involved in affective, cognitive, and somatosensory–motor tasks33.Interestingly enough, some of these brain areas, such as bilateral midcingulate cortex, left posterior and right anterior insula and left amygdala, are recognized as core regions of the central control of autonomic systemWe hypothesized that the OMT, as compared to the control condition, would induce a decrease in rCBF in a widespread network of brain regions, including (but not exclusively) those of the pain matrix and an increase in parasympathetic activity as measured by HRV.2 = 0.24, p = 0.62).Thirty-two right-handed patients were randomised and divided into the study N = 16) and control group (N = 16). Two patients (1 in the study group and 1 in the control group) dropped out during the study, leading to a final sample size of 30 patients Figure . At the 6 and conAt the end of the study period, the pain scales showed significant differences between the two groups Table .Table The control contrast T0_OMT vs T0_SHAM revealed no significant differences between the two groups for rCBF values at baseline.The whole brain analysis Fig.  showed dConsidering the region of interest (ROI) analysis, the mixed effect regression (MER) analysis showed that, as compared with control group, patients that received OMT demonstrated decreased regional CBF in the L-pINS, L-ACC, L-MFL, L-THAL and the L-CU Fig. .Figure 2Conversely, rCBF was increased in the L-LN, R-LN, L-PCC, L-SPL, R-vaINS, R-daINS, R-vplTHAL, R-OFC, R-ACC, R-MOFG and L-Cereb (CrI) in OMT patients as compared with controls Table .Post-hoc analysis revealed that these changes were statistically significant only at T2 and not immediately after the first treatment session.Table Considering regions in which CBF changes due to treatment were significantly different between groups, 6 ROIs showed a significant correlation of CBF variation with variation of VAS pain score  Table . This siSpecifically, baseline CBF changes at T2 in the OMT group were significantly correlated with the corresponding changes in VAS scores in the L-pINS , L-ACC , L-MFL , R-LN , L-PCC  and L-thalamus    but not at T1.MER showed a statistically significant difference between groups on the nuHF (01) Fig. . Tukey pSimilarly, statistically significant differences were revealed for nuLF (p < 0.01), LH/HF ratio (p < 0.01) and DFA-a1 (p < 0.05). Tukey post-hoc analysis demonstrated that in all the HRV parameters the OMT group had a significant effect compared to sham at T2 but not at T1.In addition, other time domain parameters Table  showed sOnly in the OMT group, a statistically significant negative correlation Fig.  was founFurthermore, the same brain regions showed opposite correlations at T2 between rCBF changes and nuLF changes: L-pINS , L-PCC , R-vaINS , R-daINS  and R-ACC .Also, data showed a positive correlation between nuHF and pain  in the OMT group but not in the control sample .Again, no statistically significant correlations were depicted at T1 between all parameters above-mentioned.Results from this trial showed that OMT has a two-fold effect: central, that is a change in the brain by modifying the CBF in some but not all the areas related to the pain matrix, and peripheral, that is an autonomic effect as measured by a change in HRV, specifically on HF and DFA-a1.To our best knowledge, this is the first experimental evidence of such combined modifications, providing suggestions that a mutual interconnection response might mainly mediate this brain-and-heart effect due to the administration of osteopathic treatment.After the OMT sessions there was a significant CBF decrease in some brain areas related to pain, that is L-pINS, L-ACC and L-thalamus, an increase of CBF values in the L-PCC, lentiform nuclei, R-vaINS, R-daINS and R-vplTHAL Figs. . In addi20 and in asymptomatic healthy subjects36. Interestingly, Tamburella and colleagues reported that a single OMT session produced an immediate blood flow changes in L-PCC and L-SPL21. The present study extends this previous work by examining effects in chronic LBP patients, over an extended period and combining HRV measurements. That is, while previous research has used CBF measurements immediately after a single OMT session and after three days, here we report not only that larger CBF changes are observed in response to a longer osteopathic treatment period (4 sessions over a 1-month treatment period) but that these effects are linked to a change in autonomic response. We interpret the accompanying decrease of CBF in pain-related areas as reflecting the positive influence of osteopathic treatment on heart rate variability and pain perception on LBP patients’ physiological state, hypothesizing a potential OMT effect on central mechanisms of endogenous pain modulation.These findings expand the results reported from previous fMRI studies in the same group of cLBP patientsFurther support of our hypothesis that OMT might act on CBF through the ANS by reducing the inflammatory milieu of the tissue comes from the analysis of the CBF values in combination with autonomic reactions.37. The insula, S1, and thalamus were previously linked to both acute and chronic pain39, and it was reported that these regions encode discriminative sensory components of acute pain in terms of both intensity and somatotopy42. Additionally, INS and S1 consistently show increased activity in chronic pain44. A 2016 meta-analysis demonstrated that in chronic pain studies the following areas were most frequently reported: bilateral INS—in particular the right posterior insula and the left insula, left inferior parietal lobule, left dorsal ACC (BA32), bilateral precentral (BA6&BA44) and postcentral gyri (BA40), right medial frontal gyrus, bilateral basal ganglia, i.e. putamen, bilateral thalamus and left anterior cerebellum32. Thus, CBF increases in several pain conditions, specifically in the pain regions. Our data showed that the use of 4 sessions of osteopathic treatment produced significant variations of baseline CBF values in some but not all the areas related to pain. This result might mean distinct regulation of the blood flow with a specificity of response, that is a decrease in L-pINS, L-ACC and left thalamus and an increase in L-PCC, right anterior insula, bilateral basal ganglia, right thalamus.Our findings are in line with previous studies demonstrating an increase in CBF on areas related to chronic pain. Indeed, the increase of CBF in pain-related areas was also observed in neuralgia pain patients and positively correlated to pain intensity33. These brain areas appear to be distinctively associated with cognitive output  or to somatosensory output (PCC) of CAN. These findings were recently confirmed and extended by Valenza et al., who identified additional regions such as the precuneus, angular gyrus, and cerebellum45.Nevertheless, these variations might be linked to significant changes in the control of the central autonomic network (CAN). Beissner and colleagues in a 2013 metanalysis focusing on CAN suggested that the prefrontal, anterior, and midcingulate, right ventral anterior insular and left posterior insular cortices are involved with sympathetic activity whereas the PCC, lateral temporal cortices, bilateral dorsal aINS with parasympathetic responses33. However, we need to consider that Beissner and colleagues33 as well as Valenza and colleagues45 summarized studies mainly based on healthy subjects, with and without CBF data, in rest and task-based paradigms. Interestingly, Chouchou et al., recently showed that stimulation of the anterior insula mainly induced a cardiovagal-mediated decrease in the heart rate, whereas stimulation of the posterior insula mostly produced a sympathetically mediated increase46. This might also be consistent with our results where the OMT increased the perfusion of the right anterior insula, reduced the perfusion of the L-pINS and increased nuHF, reflecting a decrease in heart rate.Therefore, we might argue that the central effect of OMT—which elicited a change in brain perfusions within specific areas also related to CAN, specifically bilateral ACC, R-vaINS, R-daINS, L-pINS, L-PCC—can be linked to the regulation of CAN on both sympathetic and parasympathetic components. Beissner et al.’s proposal on the distinction between sympathetic and parasympathetic might explain the different behaviours we found on CBF reactions in the current study47. This model was further developed recently by Smith and colleagues who better explored and contextualized the NVI model proposing eight neurofunctional hierarchical levels of control over the vagus nerve .Thayer and colleagues originally proposed the association between CAN and HRV within the Neurovisceral Integration Model (NVI), a model where the variability of heart rate, as measured by HRV, is considered a valuable proxy for the brain stateIn particular, relevant for this discussion about the role of vagus nerve as parasympathetic output and central autonomic control, is the level 6, which includes INS and ACC, that regulates processes incorporating perceptual representations of one’s current bodily state, and level 7, which includes the PCC, that further regulates these processes embracing conceptual interpretations of the meaning of both exteroceptive and interoceptive sensory input. In sum, these higher-order brain areas hierarchically regulate the neurovisceral system by, among others, controlling the cardiac autonomic output. Therefore, the fact that we observed cardiac autonomic effects on chronic pain population and that these sequelae are associated to central patterns generates discussion on (i) changes in the cross-organs relationship, more specifically brain and heart, after OMT, (ii) the relationship between brain, heart and peripheral tissue.50. Thus, modulating the cardiac output by a parasympathetic effect produces a direct effect on cerebral activity. Our data showed that a sympathetic response is associated with a decrease of CBF in some brain areas related to the pain matrix. This means that a relative reduction of blood flow might be arguably produced based on a sympathetic response. This might be conceptually understood as pain patients express higher baseline CBF values in these regions compared to asymptomatic-healthy subjects.Interestingly, examples of coordinated cross-organ reactions include the impact of cardiac functions on cerebral activity54, acting as a neurogenic modulator when hemodynamic changes challenge cerebral activity. Therefore, applying a treatment, which produces a reduction of pain, might justify lower perfusion of these areas and thus a relative decrease of blood flow51. On the contrary, exploring the cardiac data, the findings of the present study are consistent with previous research on adults, which concluded that osteopathic approach produces an increase in parasympathetic tone leading to a trophotropic effect12. Thus, a double effect seems to be produced where a prevailing sympathetic response is observed centrally, whereas a major parasympathetic effect is showed peripherally.Clinical and laboratory research demonstrated the role of sympathetic branch in preventing increase of CBF13. While we did not include a measure of pro-inflammatory levels in this study, it has previously been reported that in a sample of LBP patients the use of OMT can reduce cytokines levels16 and this reduction is associated to a reduction of pain perception15. Our sample showed a significant reduction of pain perception and better overall clinical conditions, possibly related to a reduction of pro-inflammatory state in the tissue leading to a change in the neurogenic neuroinflammation55. The ANS, CAN and interoception mediate all these processes. Modifying the metabolic state of the tissue, therefore, produces a proxy for unmyelinated c-fibers that in turn, through the spinothalamic tract, results into the activation of the insular cortex, specifically the posterior lobe of the left insula59.To further explore this hypothesis, it has been argued that the osteopathic approach might act through an anti-inflammatory and hyperparasympathetic effect mediated by interoception58, where the insula receives input from other areas of the brain including hedonic areas . Therefore, we might argue that the use of osteopathic treatment might produce a significant change in the cerebral activity, specifically on key areas related to CAN and thus to interoception and interoceptive control.Interestingly, our data showed that the left posterior INS is a specific target of the osteopathic effect. Also, the other areas involved after OMT are consistent with the hypothesis of the awareness model60. Therefore, comparing immediate effects with long-term outcomes would shed light on the brain changes that might occur. Furthermore, although the post-hoc analysis showed an adequate power for the majority of the ROIs, future CBF studies might benefit from an a priori power calculation.The present study only examined the immediate and 1-month sustained effect of OMT on chronic pain patients. Future work is needed to determine whether there are longer-term, clinically and brain significant, benefits of this type of manual intervention and if so, what dose, in terms of frequency and duration, is required. Systematic evaluation of the effects of OMT on chronic pain patients by Cerritelli and colleagues have determined that eight treatment sessions over a 6 month period are sufficient to decrease pain, disability and migraine attacks per monthIn conclusion, osteopathic interventions can have some clinical benefits, but a greater understanding of the neurobiological mechanisms is needed to optimize pain protocols and ameliorate the long-term negative consequences of pain. The present study supports the hypothesis that OMT can induce CBF changes by acting through autonomic responses, offering insight into the development and optimization of novel pain care strategies.The study was designed as randomised placebo-controlled to explore possible mechanisms to explain the effect of osteopathic manipulative treatment compared to sham treatment on cerebral blood flow in a sample of cLBP patients.61. Exclusion criteria were: clinical sign of neurological damage with sensorimotor impairments ; suspected or confirmed spinal pathology ; history of spinal surgery (e.g. decompensation or stiffening); whiplash incidence within the last 12 months; cervical pain that reduces active movement to less than 30° rotation to each side; known vestibular pathologies; major surgery scheduled during treatment; physiotherapy during the last 12 weeks; inability to follow the procedures of the study: e.g. due to language problems, psychological/psychiatric disorders, dementia of the participant; parallel participation in another study. At enrolment, eligible patients were assessed by a senior MD to confirm the diagnosis and exclude psychiatric disease and/or any other exclusion criteria.Adult patients with chronic Low Back Pain were recruited from a rehabilitation centre of the university of Chieti (CUMFER). Inclusion criteria included any chronic (> 3 months) pain or discomfort localised below the costal margin and above the inferior gluteal folds, with or without referred leg painEligible patients were randomly divided into two groups using a 1:1 ratio and assigned to the study (OMT) group or the sham group. Block randomization was applied according to a computer-generated randomization list using a block size of 10. All patients were not aware of any step of the study design as well as outcomes or group allocation. The randomization list was stored in a dedicated and protected web-based space and an information technology consultant was in charge for the entire process.Research staff were unaware of the study design and outcomes. Moreover, they were blinded to patients’ allocation, since all patients were touched by the practitioner. Only the osteopath was aware of the patients’ allocation. Moreover, the practitioner who performed OMT had no role in patient care decisions. Researcher using the MRI and dealing with MRI data were unaware of patients’ allocation.The study was approved by the local ethics committee (University of Chieti-Pescara number: 7/09-04-15) and all subjects provided informed written consent, in line with the Declaration of Helsinki. The current protocol was registered on clinicaltrial.gov (ID: NCT02464475) on 08/06/2015. All methods were carried out in accordance with relevant guidelines  and regulations.Before the MRI session, patients were asked to complete paper-based questionnaires. A socio-demographic questionnaire was used to collect baseline data.62. The State-Trait Anxiety Inventory (STAI-Y1 and Y2) was used to test trait anxiety63.The Edinburgh Handedness Inventory was utilized to investigate the hand dominance66.Furthermore, the TEMPS-A  was administered to measure the affective temperament that defines the bipolar spectrum, with depressive (D), cyclothymic (C), hyperthymic (H), irritable (I), and anxious (A) subscales68. Questionnaires were administered at enrolment, at each session and at the end of the study period.Several tools were specifically used to assess pain perception in patients: Visual Analogue Scale (VAS), the Roland–Morris Disability Questionnaire and the Oswestry Low Back Disability Questionnaire (OSW) for the physical disability due to low back pain69. Osteopathy is based on the structural assessment to diagnose somatic dysfunctions and a series of manipulative techniques for the treatment70.Patients eligible for the study were divided and randomised in an OMT and Sham group. Patients allocated to the OMT group received 4 osteopathic sessions (approximately 30 min each). Osteopathy is a drug-free manual medicine, where osteopathic practitioners use manual techniques to diagnose and treat somatic (body framework) dysfunction (ICD-10 code: M99.0–99.9)In the present research the treatment was administered by a licensed and registered osteopath. Among the series of techniques, the techniques used in the current study were balanced-ligamentous, balanced-membranous and fluidic techniques, in line with the principles and procedures available in the current osteopathic literature. All treatment sessions took place in the CUMFER.The sham group received an osteopathic-like manual assessment and treatment, that means the practitioner applies a manual contact without using any type of specific osteopathic technique or procedure. After the evaluation, the operator asked the patient to lay down on the plinth and gently placed the hands on a pre-defined set of bodily parts without applying any type of technique but just using a gentle static or dynamic touch. The parts identified in the protocol were: low back, sacrum, pelvis, diaphragm, upper thorax, cervical spine and cranium. The sequence to apply during the session was decided by the operator before the session. This was planned to prevent any possible chance from the patient to guess the group allocation. The sessions lasted 30 min, as for the OMT, took place in the same location/room and were administered by the same practitioner. Besides, the operator was instructed to maintain and establish the same type of patient-doctor relationship. This procedure was used to avoid any possible contamination and to prevent allocation bias. Patients assigned to the sham group received osteopathic treatment following completion of the trial.During the study period, all patients were asked to avoid drug consumption.20, the 3 time points were established : before the treatmentImmediate response (T1): Immediately after the first manual sessionSustained response (T2): at the end of the study period (after a month), which included 4 treatment sessionsAs for a previous BOLD studyAfter the clinical evaluation (enrolment) and at T2, patients were asked to fill in the paper-based questionnaires.71 with the labeling parameters optimized according to a recent white paper72: postlabel delay 1900 ms, label duration 1750 ms. Background suppression pulses were used at 2110 ms and 3260 ms after start of labeling. Other imaging parameters were TR/TE = 4269/10 ms, SENSE factor 2.3, matrix 64 × 64, voxel size 3.6 mm × 3.6 mm × 5 mm, 19 slices acquired in ascending order, 60 dynamics. The labelling plane was positioned 85 mm below the AC-PC line72. An equilibrium magnetization image (M0) with a long TR  was also acquired for calibration purposes using readout parameters identical to the pCASL sequence. The total duration of the pCASL measurement was about 5 min.MRI was performed using a Philips Achieva 3 T scanner . A whole-body radiofrequency coil for signal excitation and an 8-channel phased-array head coil for signal reception were used. A high-resolution structural volume was acquired using a 3D fast field echo T1-weighted sequence . Then, Blood Oxygen Level Dependent (BOLD) fMRI data were acquired. Perfusion imaging was performed using a pseudo-continuous ASL (pCASL) sequence20. Importantly, we carefully controlled potential confounding factors requiring the subjects to refrain from caffeine, nicotine or alcohol consumption for at least 6 h before the MRI exam. In addition, the period within the day where the acquisition was performed, and the time spent in the scanner before the ASL measurement was the same for each subject and session to exclude effects due to circadian cycle, fatigue and menstrual cycle.The MRI session also included the acquisition of BOLD data, part of which were recently published73.During all MRI measurements, respiratory and cardiac physiological activity was recorded using the built-in scanner devices. Specifically, respiratory cycle was recorded with a pneumatic belt strapped around the upper abdomen, whereas the cardiac cycle was registered with a pulse oximeter placed on a finger of the right hand. Both signals were acquired during the pCASL sequence and sampled at 100 Hz, allowing a reliable assessment of HRVIn addition, subjects were instructed to stay awake with their eyes closed and minds clear. Maximum care was taken to avoid situations that may trigger abnormal pain. Subjects laid in a supine position on the scanner table with head immobilized by foam padding.74. First, motion correction was performed to realign all tag/control time frames to a reference image represented by the first tag/control volume. Second, an additional coregistration matrix between these two reference images was determined to account for possible head motion between the two acquisitions, using a mutual information-based approach75. These preprocessed control/tag timeseries were coregistered to the M0 image that was also used to determine a coregistration matrix between perfusion images and the structural scan. To take into account intrasubject alignment across the three sessions, the structural scan at T0 was used as the reference image for all coregistrations. Finally, individual subject data were normalized to the MNI space using linear and nonlinear transformations (AFNI script “@SSwarper”).Analysis of pCASL data was carried out using AFNI. Preprocessing was performed on the tag and control ASL images separately76:Voxel-wise quantitative CBF maps were derived using a single compartment model72, αinv = 0.83 is a correction factor for the background suppression77, M0 is the equilibrium magnetization signal, τ is the label duration (1750 ms) and λ is the blood–brain partition coefficient (0.9 ml/g).Here, SIC and SIT are the time averaged signals of control and tag images respectively, PLD is the post label delay , T1A is the longitudinal relaxation time of arterial blood (1650 ms at 3 T), α = 0.85 is the labeling efficiencyThe calculated CBF maps were then normalized to the MNI space using the previously determined spatial transformations.p < 0.01 corrected for multiple comparisons using False Discovery Rate (FDR).The following contrasts between voxel-wise CBF maps were considered: (a) (T1_OMT–T0_OMT) vs (T1_SHAM–T0_SHAM) describing the post first treatment vs. baseline, (b) (T2_OMT–T0_OMT) vs (T2_SHAM–T0_SHAM) showing the end of study period vs. baseline, (c) (T2_OMT–T1_OMT) vs (T2_SHAM–T1_SHAM) defining the end of study period vs. post first treatment. These contrasts represent treatment’s effects controlled by baseline, i.e. T0, scans, and delayed effect, respectively. The additional contrast T0_OMT vs T0_SHAM was performed to control for potential differences between groups at baseline. The group statistical maps obtained from these contrasts were thresholded at http://kubios.uef.fi) to compute HRV parameters. Standard 5-min recordings were considered for RR series. There were no significant differences in the percentage of artifactual epochs identified between the two groups.Standard 5 min recording (corresponding to the pCASL acquisition) was used to derive HRV data. Inter-beat intervals were extracted from the pulse oximeter. Using customized software , outliers, i.e. artefacts and ectopic peaks, were identified and removed from the data. Specifically, to account for non-experimental movements that might produce extreme values, individual participant’s datapoints that were more than three S.D.s above or below the whole sample mean were identified. Such datapoints were determined to be additional artefacts and replaced by the mean of that participant’s non-artifactual epochs. The artifact percentage accepted for each individual was set at 2%. Two participants (1 in the OMT group and 1 in the SHAM group) were identified as having artifactual epochs, corresponding to 6.25% of period datapoints across participants. Intervals were then imported in Kubios software  Standard analysis , and (2) Time-varying analysis. For time-domain analysis methods, the window width of the moving average filter was set at 5 beats. Also, the threshold used in the computation of NNxx and pNNxx parameters was set at 50 ms, thus NN50 and pNN50). From power spectra (Fast Fourier transformation -FFT- using Blackman Harris window) of equidistant linear interpolated (4 Hz) tachograms (resampled to 2 Hz), the following frequency domain standard HRV indices were used for linear analysis: nuHF, from 0.15 to 0.4 Hz, i.e. signal of parasympathetic heart rate modulation80; nuLF, from 0.04 to 0.15 Hz, i.e. predictor of sympathetic modulation81; LF/HF ratio as being demonstrated a valid marker of ANS activity82. In addition, the spectrum estimation option was 300 points/Hz, the FFT spectrum using Welch’s periodogram method has a window width of 300 s and a window overlap of 50%. The AR spectrum used an AR model order of 16. The nonlinear analysis options included embedding dimension at 2 beats with a tolerance of 0.2 times SD. The detrended fluctuation analysis used a short-term parameter ranging between 4 and 12 beats and long-term fluctuations between 13 and 64 beats. Considering non-linear analysis, the DFAα1 parameter was computed. DFAα1 is considered a sensitive parasympathetic index83 able to discriminate possible long-term correlations and complexity of RR interval series84. A fractal structure of heart rate was quantified by estimating a short-term, alpha 1, fluctuations, obtained from the range 4 ≤ n ≤ 16. The time-varying analysis settings were: HRV analysis window width equal to 300 s with an effective data threshold at 50%. For the time-varying spectrum estimation, the well-known spectrogram was chosen.HRV analysis method, based on processing recorded RR intervals, was divided into linear analysis (time and frequency domain) and nonlinear analysisd, alfa = 0.05 and number of subjects equal to 16.Due to lack of perfusion studies assessing the effects of manual therapies on cLBP, the computation of a priori sample size was difficult. For this reason, we computed a posteriori the effect size (Cohen’s d) for each ROI by using the baseline-T2 difference data in contrast between OMT and sham group. In addition, β values were calculated using pwr.t.test function {pwr} in R considering Cohen’s Arithmetic mean and standard deviation as well as median, percentage and range were used to explore the general characteristics of the study population. To compare the OMT and sham group at enrolment, univariate statistical tests, student t test and chi square test were performed.85, over M patterns. Shapiro test was applied to test the normality of R-R distribution (p < 0.05). In case of non-normal distribution, a log transformation was applied. Then, M patterns were assessed for normality. The patterns were randomly selected from a set of sequences of length L85. These patterns were then used for the final analysis.HRV analysis was performed in the present trial using the restricted weak stationarity (RWS) test to assess stationarityTo study the independent effect of osteopathic manipulative treatment on perfusion data, HRV endpoints and pain data, a 2 × 3 repeated measure analysis based on MER model considering random effect for groups (OMT-SHAM) and a fixed effect for period (T0–T1–T2) was used to further explore any difference. The following confounding factors were considered in the analysis: age, gender, caffeine consumption, circadian and menstrual cycle. Post hoc pairwise analysis adjusted by Holm–Bonferroni correction was utilised after any statistical difference resulted from MER.To indicate statistical difference, two-tailed P values of less than 0.05 was considered. This data analysis was carried out using the R statistical program (v. 3.5.2).To determine whether CBF in the pain-related regions was correlated with pain intensity, Pearson correlation analyses between regional CBF values and VAS scores at the different timepoints were performed. Further Pearson-based correlation analyses were conducted exploring correlation between rCBF and HRV parameters as well as between HRV and pain.86, ROIs were defined as spherical nodes (6 mm of radius) using independent coordinates from the literature45 (Supplementary materials—Table Bonferroni correction for multiple comparison was used. In order to avoid circularity problems in the analysisThese ROIs showed a good spatial overlap with the pain-related regions revealed by the voxel-wise contrasts in the above analysis.Supplementary Figure 1.Supplementary Figure 2.Supplementary Information.


To investigate the effects of sesame oil extracted from tahini (Ardeh) on pain severity in patients with upper or lower limbs trauma. st through November 30th, 2016. The patients were randomly assigned to two groups using block randomization. The intervention group received topical sesame oil extracted from tahini (Ardeh) and the placebo group received cooking oil. Pain severity, pain sensitivity and heaviness of painful site were assessed. This double-blinded randomized clinical trial study was conducted on 120 patients with upper or lower trauma in Shahid Rajaee Hospital, Shiraz, Iran, from May the 1Overall, we included 90 patients with traumatic limb injuries in this study who were randomized to two study groups. The mean age of the patients was 28.3 ± 6.8 (ranging from 25 to 35) years and there were 63 (70%) men and 27 (30%) women among the patients. In the sesame oil group, the mean changes in the pain severity , pain sensitivity  and heaviness of painful site  were significantly lower when compared to the placebo group in the second day of the intervention. None of the patients experience adverse drug effects. Our findings suggest that the topical use of sesame oil extracted from Tahini has a pain reliever effect on the skin after bruising and it helps prevent skin discoloration in patients with traumatic injuries of limbs.  A bruise is the result of smaller vessels’ bleeding into subcutaneous tissues , 2. LeakSesame is one of the most ancient crops cultivated which has been used for more than 5 thousand years in India ; amongstSesame paste  is a complex colloidal mixture among the family of oils, which is a rich source of protein and lipid. De-hulled sesame seeds are milled and roasted without any changes in its component to make this material . This maStudy populationThis double-blinded randomized placebo-controlled clinical trial study was conducted on 120 patients with upper or lower limbs trauma referred to emergency department of Shahid Rajaee Hospital, Shiraz, Iran, from May the 1st through November the 30th, 2016. All patients with one or multiple blunt trauma in the upper extremity  or lower extremity  occurring at least 1 hour and maximally 6 hours after admission were enrolled in the study. Inclusion criteria were age range of 15- 40 years, lack of any sign of bone fractures, internal or external bleeding, dislocation, amputation, presence of a foreign body, nerve damage, fever as well as infection, lack of cast or splint at the trauma site, lack of regional pain based on Visual Analog Scale (VAS), lack of any history of addiction, cigarette and alcohol abuse, lack of any sensitivity or allergy to the sesame plant group, not receiving drugs or herbal extracts which may interact with the study therapeutic protocol such as anti-coagulant and analgesics, and lack of diseases that may affect the pain severity such as diabetes, cardiovascular, liver, kidney, and musculoskeletal diseases. Exclusion criteria were having any sign of allergy to the sesame oil or peptic or duodenal ulcers, receiving treatment out of the study, inappropriate follow up by patients (missing follow-up more than two times), and patient’s willingness to withdraw in any phase of the study.Ethics and blinding This study was approved by the ethics committee of International Branch of Shiraz University of Medical Sciences, Iran, with the code No. IR.SUMS.REC.1394.34 and registered in the Iranian Registry of Clinical Trials (IRCT) with the code No. IRCT20171017036838N1. Before starting the interventions on patients, the researcher obtained the informed consent, verbally explained about the research, and assured the patients of confidentiality and anonymity of their data. Apart from the project coordinator, the patients, the staff involved at the clinical center, and members involved in collecting and analyzing the data were blinded to the intervention allocation. During the intervention, neither the patients nor the researcher knew which containers were treatment and placebo.Randomization and interventionPatients were randomly allocated to intervention and control groups using block randomization method. They generated 30 randomization sequences using computer-generated random numbers in permuted blocks of size four. Both groups received a 5 min light massage on the trauma site three times of a day with 8 h interval for 2 days. Patients in the intervention group received topical sesame oil extracted from tahini (Ardeh), whereas the patients in the control group just received cooking oil with the same color and container to avoid bias in the results. Also, routine cares were implemented based on the care protocol for all patients with trauma. According to this protocol, the periphery and the center of the trauma site were irrigated and cleaned with 1000 mL sterile normal saline solution (0.9%), and then dried with a sterile pad and massaged. Furthermore, based on our pilot study conducted on 10 patients in each group (not included in main sample), the topical usage of both oils showed no adverse effects. 10 drops (about 1 mm thick and 3.8 mL) of each oil extract was applied for each 50 square cm on the site of trauma, using a dropper. After that, the site of trauma was massaged lightly. Study protocol At initial meeting considered as baseline, pain severity, pain sensitivity and the level of painful site heaviness, in addition to demographical data and the background of the NSAIDs received were assessed for both groups by the same trained specialist. Then, all the patients were taught how to take care of their trauma during 2 h in the consultation room at the recruitment hospital. All the patients were educated to put cold compress on the trauma site on the first day, put warm compress on the following days, and avoid usage of other medicines or herbal extracts for pain relief. The researcher assessed the characteristics including age and gender. For measuring the pain severity, we used Visual Analogue Scale (VAS). When responding to a VAS item, the respondents specified their level of agreement to a statement by indicating a position along a continuous line between the two end-points (0 and 10). Two other scales including pain sensitivity and heaviness of painful site were also measured. Like VAS, these two scales were measured by a continuous line between the two end-points (0 and 10) . The datSample SizeTo estimate the sample size, we used the result of a previous study  which inStatistical Analysisp-value less than 0.05 was considered as statistically significance.Statistical analysis was performed using the Statistical Package for Social Sciences . Descriptive statistical tests, mean, standard deviation frequency and percentage were used for demographic characteristics . IndepenOverall, 120 patients with inclusion criteria participated in the study, where 10 patients were excluded during the intervention period due to NSAIDs consumption and 20 lost the follow up. Of the remaining 90 patients, 49 (54.4%) were in the sesame oil group and the rest of them in the placebo group (45.6%). The flow diagram of the study is demonstrated in p<0.001). In addition, comparison of the mean differences of the baseline scores and those after 48 hours between the placebo and intervention groups showed a significant difference (The mean age of the patients was 28.3 ± 6.8 (ranging from 25 to 35) years and there were 63 (70%) men and 27 (30%) women among the patients. Both study groups were comparable regarding the baseline characteristics and the demographics . In ordefference . Figure et al.’s study, massage with topical sesame oil led to a significant reduction in pain severity of patients with limbs trauma. This study finding showed that on the 9th day of intervention, pain severity decreased 81% in the sesame oil group, while it decreased only 18% in the placebo group compared to the baseline (first day) [et al. and Hirsch et al.’s studies, the effect of an ointment containing sesame oil in combination with other herbal extracts and beta-Sitosterol on patients with burn was evaluated. It was shown that massage with moist exposed burn ointment (MEBO) decreased pain in these patients [According to the results, sesame oil extracted from Tahini led to a significant decrease in pain severity, pain sensitivity and heaviness of painful site on the second day in the intervention group compared to the placebo one. The results also showed that in the placebo group the scores of pain severity and pain sensitivity changed significantly in the negative directions. Along with the pain relief property of sesame oil, we observed that this substance prevented skin discoloration due to bruising. In the literature review, we could only find two similar studies that evaluated the effect of sesame oil on trauma. Based on Nasiri rst day) . The othrst day) . Althougrst day) -35. In Apatients , 34. Zahpatients . In anotpatients .On the other hand, there were some animal studies that pointed to the analgesic properties of sesame oil  in reducing pain due to different causes -42. For Although the high content of unsaturated fatty acids , lignans  and gamma-tocopherol responsible for pain relief are sesame oil benefits , the exaIn conclusion, our findings suggest that the topical use of sesame oil extracted from Tahini has a pain reliever effect on the skin after bruising and it helps prevent skin discoloration. Further larger studies are required to confirm the results of the current study.


Patient‐controlled subcutaneous analgesia (PCSA) with sufentanil is an alternative analgesia strategy in patients with stage III‐IV cancer; however, its efficacy and safety have not been fully investigated.From May 10, 2017 to November 10, 2017, 120 patients with stage III‐IV cancer suffering from moderate to severe pain were prospectively enrolled from six hospitals and randomized to receive PCSA with morphine (control group) or sufentanil (intervention group). Before the PCSA and on days 1, 3, 7, 14, 28, and 56 after treatment, the numeric rating scale (NRS) and 36‐item Short Form health survey (SF‐36) were completed for each patient and the side effects were also recorded.P > .05) were observed in the preoperative NRS score and the SF‐36 parameters between the two groups. Patients in the intervention group achieved better pain relief, as indicated by lower NRS scores at days 14 (P = .040), 28 (P < .001), and 56 (P < .001) after PCSA device implantation (vs control group). Furthermore, the patients in the intervention group also achieved a better life quality, as indicated by the physical role, general health, social function body pain, and mental health scores. Finally, the patients receiving sufentanil showed lower levels of nausea and somnolence than those in the control group.No significant differences (PCSA with sufentanil achieves better pain control and life quality as well as fewer adverse reactions in stage III‐IV cancer patients with pain and may be a promising pain management in these patients.This study was registered at chictr.org.cn with the trial number: ChiCTR‐IPR‐17011280. PCSA with opioid or sufentainil in home care could effectively relieve stage III‐IV cancer pain. Sufentainil PCSA could provide a better analgesia, improved life quality and less adverse events than morphine. Currently, the worldwide pain prevalence is about 50% in cancer patients at the time of diagnosis and approximately 80% in patients with stage III‐IV cancers.In the clinical setting, rapid‐onset analgesia can be achieved through a variety of methods, including subcutaneous (s.c.) or intravenous (iv) patient‐controlled analgesia (PCA),Sufentanil, a highly lipophilic opioid fentanyl analogue, is typically used for surgical analgesia, especially in patients with renal impairment, based on pharmacokinetics and clinical experience.Thus, the primary purpose of the current study is to prospectively evaluate the efficacy of patient‐controlled subcutaneous analgesia (PCSA) with sufentanil for home care stage III‐IV cancer patients. The secondary purpose is to evaluate the quality of life in these advanced cancer patients.22.1This prospective randomized controlled clinical trial was approved by the Ethics Committee of the First Affiliated Hospital to China Medical University No: 2016‐175‐2) and registered at chictr.org.cn with the number of ChiCTR‐IPR‐17011280. All stage III‐IV cancer patients were prospectively enrolled from six research centers from May 10, 2017 to November 10, 2017  cancer patients of 18‐year‐old or above; (2) patients with a numeric rating scale (NRS) pain score ≥ 5; Karnofsky performance score (KPS) ≥40; (3) patients with advanced cancer pain cannot take oral analgesics; or take oral morphine at a dose over 100 mg/d without good pain control; or cannot tolerate opioid side effects; and (4) patients voluntarily participated in this study with signed informed consent.Patients were excluded for the following reasons: (1) refusal to participate in this trial; (2) poor general situation, unable to describe the symptoms; (3) allergy to the medications; (4) inability to tolerate treatment; (5) consciousness disorder; (6) severe infection or respiratory insufficiency; (7) contraindications to test drugs; (8) history of drug abuse; and (9) pregnant, lactating women; or subjects who have a pregnancy plan within 1 month of the trial.2.3According to a computer‐generated random allocation sequence, a total of 120 patients were enrolled and randomly assigned into one of two groups: the intervention group, in which patients received PCSA with sufentainil (n = 60), and a control group, in which patients received PCSA with morphine  was implanted into the subcutaneous tissue around the navel and the PCA pump was connected After randomization, the research associates collected patient information including age, gender, weight, location, and duration of pain. Before the initial treatment (baseline) and on days 1, 3, 7, 14, 28, and 56 post‐treatment, the research associates asked patients to rate their NRS scores. The adverse reactions, including bleeding, infection of the puncture site, subcutaneous induration, nausea, vomiting, urinary retention, and somnolence, were also recorded. All patients were followed up once a week to replace the quick‐set catheter. Additionally, telephone follow‐ups were performed every 3 days to evaluate any adverse reactions. Patients or their family members were able to phone the doctor in case of any emergency.The initial daily dose of morphine depended on the prior pain treatment. Briefly, the 1/3 dose of oral opioid during the previous 24 hours was given for severe opioid tolerant patients. If the patient suffered from moderate pain (rest NRS score ≧4), a 50% increase in the initial dose was administered.2.4Data were collected at baseline (1 day before treatment) and in the morning (8:00‐10:00) on days 1, 3, 7, 14, 28, and 56 after the PCSA procedure. The patient data were excluded from analysis if the patient died within the 8‐week follow‐up period.The NRS scores were evaluated at 8, 16, and 24 o'clock of every measuring day and the average scores were calculated. The Chinese version of the Short Form health survey (SF‐36) 2.5On the observation of bleeding or infection at the puncture site, the implanted catheter was removed immediately and local antimicrobial treatment was delivered. A new puncture point was located at least 10 cm away from original puncture point. The data of patients with bleeding or infection were collected but not used for the analysis, though they were analyzed based on the intention‐to‐treat principle. A magnesium sulfate solution was used to compress subcutaneous induration. Nausea, vomiting, urinary retention, and somnolence/drowsiness were treated using conventional therapies.2.6According to previous literature, approximately 70% of patients with cancer suffer from moderate to severe pain.2.7P value < 0.05 was considered to be statistically significant.The normal distribution of the continuous variables was tested using the Kolmogorov‐Smirnov test and all measurements fell in normal distribution. Values are expressed as mean and 95% confidence interval (CI). Categorical variables were analyzed with Fisher's Exact Test. Statistical analysis was performed using the Statistical Package for Social Sciences version 19.0 . A 3A total of 163 patients were initially enrolled in this study and 43 patients were excluded for the following reasons: refusal to participate in the study (n = 25), poor general situation (n = 13), and other reasons (n = 5). The demographic characteristics of the patients, including age, gender, body weight, disease duration, and pain score before treatment, were similar between the two groups . The intervention induced better pain relief indicated by the significantly decreased NRS scores in the intervention group than the control group at 14 , 28 , and 56  days after pump implantation  days after pump implantation , 3 , 7 , 14 , 28 , and 56  days after pump implantation , 28 , and 56  days after pump implantation , 28 , and 56  days after pump implantation  days after pump implantation , 3 , 14 , and 56  days after pump implantation  except for the nausea  and somnolence  and PCSA are two common techniques that are used to maintain drug concentrations to be stable enough to meet the analgesia requirement of patients. Terminal cancer patients in China cannot access required medical resources due to the limited hospital resources and severe financial burden. We aimed to obtain better and safer pain relief for patients with stage III‐IV cancer. In agreement with previous reports, in the current study, PCSA sufentanil provided comparable or improved rapid onset analgesia compared to that with morphine. Meanwhile, PCSA sufentanil provided a better life quality and fewer adverse events than that with morphine, which is consistent with a small population pilot study in 1995.No severe adverse event was observed in the current study. The likely reasons are as the follows: (1) PCSA is a safe drug delivery technique, as reported in a previous study.This study has limitations. First, this study was performed in only one province and further multi‐site investigations are necessary to validate the conclusion. Second, it was difficult to recruit a large number of patients because 26.4% of the eligible patients refused to participate, due to their unwillingness to undergo PCSA, which may have led to the selection bias.5PCSA with opioid or sufentainil in home care could effectively relieve stage III‐IV cancer pain. Sufentainil PCSA could provide a better analgesia, improved life quality, and less adverse events than morphine.The authors declare that they have no conflict of interests.Cheng‐Fu Wan: clinical experimental design, clinical trial practice; statistical analysis of data and manuscript writing; Qing‐Zhu Meng: clinical trial practice; Yan‐Wei Wang: clinical trial practice; Liang Qi: clinical trial practice; Chang‐Liang Ai: clinical trial practice; Xin Sui: clinical trial practice; Tao Song: clinical experimental design and manuscript review.This prospective randomized controlled clinical trial was approved by the Ethics Committee of the First Affiliated Hospital to China Medical University (No: 2016‐175‐2) and registered at chictr.org.cn with the number of ChiCTR‐IPR‐17011280. All procedures performed in studies involving human participants were in accordance with the ethics standards of the institutional and national research committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethics standards.


Cancer induced bone pain (CIBP) is frequent in patients with non-small cell lung cancer (NSCLC). Radiation therapy continues to be the gold standard for treatment of painful bone metastases, however only a limited number of metastases can be irradiated. We evaluated non-radiation based early CIBP relief options in NSCLC through a systematic review.Systematic review including all prospective articles published between 01-1994 and 06-2020 on Pubmed, Cochrane Library and ClinicalTrials.gov database. Inclusion: non-radiation based trials evaluating CIBP early pain relief options  in ≥10 NSCLC patients. Radioisotope trials were excluded as these treatments have interactions with systemic anticancer therapy.188 articles were found; 10 articles (6 randomized controlled (4 double blinded), 1 phase II single-arm, and 3 prospective trials) fulfilled the inclusion criteria. Six of these trials consisted of ≥2 treatment arms, whereas the others were single-arm studies. In total, 554 NSCLC patients were evaluated in these trials. The included trials were very heterogeneous regarding evaluated treatment options, methods of pain measuring, and endpoints. No high-level evidence for specific early pain relief treatment options was found.Non-radiation based studies evaluating treatment options to rapidly reduce CIBP in NSCLC are scarce. This systematic review shows that there is no high-level evidence to recommend a specific treatment for early pain relief. Future research should focus on early pain relief treatment options for CIBP in NSCLC. Bone metastases are diagnosed in 24%–70% of patients with non-small cell lung cancer (NSCLC) during the course of the disease –3. Up toTumor invasion into bone causes osteoclast and osteoblast recruitment and modulation of genes and proteins involved in the bone microenvironment. Numerous factors are involved in the process of bone pain such as nociceptor activation and sensitization, ectopic sprouting of nerve fibers and central sensitization in the spinal cord and brain. Without treatment of the underlying disease and/or local treatment, no bone healing occurs in bone metastases, leading to a vicious circle of CIBP, central sensitization resulting in more pain, and the development of chronic bone pain . TherefoDue to the high incidence, chronic character, and negative impact on QoL and OS, CIBP is an important issue that needs to be addressed in metastatic NSCLC. According to the World Health Organization (WHO) pain ladder, (bone) pain should first be treated with paracetamol and/or non-steroidal anti-inflammatory drugs (NSAIDs), followed if necessary by adding mild, and later strong opioids . The extThe European Society for Medical Oncology (ESMO) guideline on metastatic NSCLC (2018) recommends denosumab or zoledronic acid in patients with NSCLC and bone metastases considered at high risk for skeletal related events (SREs) and with a life expectancy of >3 months  . This reThe ESMO guideline on bone health in patients with cancer (2020) states that multidisciplinary management  is needed for effective treatment of metastatic bone disease. They suggest radiotherapy as treatment of choice in localized CIBP, but no specific treatment recommendations are made for diffuse CIBP . The NatSurvival is improving in patients with NSCLC, partly because of the survival benefit seen with immune checkpoint inhibitors for a large proportion of patients and partly due to the availability of tyrosine kinase inhibitors for the group of patients with an oncogenic driver. As it is possible that these patients live longer with CIBP, effective, preferably early pain reducing treatments might be more relevant. We performed a systematic review specifically focusing on early non-radiation based pain relief options for NSCLC patients with CIBP.A systematic search of the literature published between January 1994 and June 2020 was performed using the PubMed, the Cochrane Library and the ClinicalTrials.gov database. Published studies were identified using a search strategy based on the Patient-Intervention-Control-Outcome (PICO) method shown in . PRISMA Two authors (AB and BB) independently screened the titles of the selected studies and subsequently the abstracts of the eligible studies. The same authors independently examined the full texts of the selected articles regarding the inclusion criteria. Studies were included if they met the eligibility criteria. To complete the search, the references of all eligible articles were manually searched for additional relevant articles. Also, the excluded review articles were screened for relevant studies which were not represented in the original search. The entire search and selection were independently checked by a third reviewer (LH). In case of disagreement during study inclusion, consensus was sought.When available and applicable, the following data were extracted from eligible studies by one author (AB) and independently by another author (BB): year of publication, number of study arms, randomization method, duration of study and follow-up, histological diagnosis, intervention , method of pain score , timing of pain score, efficacy of intervention on pain relief, whether results were specifically for NSCLC or for all included patients, and primary and secondary objectives of the trials. Final approval of the extracted data was performed by LH.The Jadad scale was used to assess the methodological quality of the included trials . We did The literature search identified 186 articles in total without duplicates. As mentioned in the inclusion criteria, reviews were excluded in the search strategy, but to broaden the search results these reviews were manually searched for relevant studies. After checking the reference list of the reviews identified with the systematic search, 2 additional relevant articles were included. Of these 188 articles, 151 were excluded because of non-relevant titles. 18 of the 37 remaining articles were excluded because they did not fulfill the inclusion criteria based on the abstract. After screening of the full text of the remaining 19 articles, 14 articles were excluded because of: no answer on the clinical question (N = 4), radiotherapy as treatment modality (N = 1), radioisotopes as treatment modality (N = 1), retrospective study or case report (N = 2), and a language barrier . After manual search of the reference list of included articles 5 other relevant articles were included. The flowchart for article selection is shown in One phase II trial , six ranThe primary objectives of the included trials varied from efficacy and safety of combined treatment of chemotherapy and zoledronic acid , 29 effeThe studies of Yoh et al. and Zarogoulidis et al. used the BPI as method of measuring pain score , 30. In Five studies  showed a significant treatment effect on pain score , 27, 29.Of note, none of the studies including patients with different primary tumor histologies reported results for the subgroup of patients with NSCLC. EGFR mutation or ALK fusion , but this resulted in limited number of eligible trials. To be more inclusive, we broadened the time of “early pain relief” to a maximum of six weeks. Even then, only ten studies were eligible. Of note, the included trials were very heterogeneous regarding treatments evaluated, primary endpoints, methods of pain measurement and timing of assessment. Importantly, the randomized trials included patients with different histologies, and patients with NSCLC only comprised a subgroup in these randomized trials (554 [44%] of included patients). Importantly, not all treatments evaluated are comparable with recommended pain treatment in clinical guidelines. For example, according to international and national guidelines for breakthrough cancer pain, shorting-acting morphine should be added to standard dose long-acting morphine to treat breakthrough pain . Three oIn most other studies , 29, 30,What are other possible treatment options for CIBP in NSCLC? The first step to achieve early pain relief in CIBP is analgesics according the WHO pain ladder . This adTo the best of our knowledge, there is no explanation why for example bisphosphonates showed a reduction of CIBP in some malignancies but not in lung cancer , 44, 45.Radioisotopes  are an alternative treatment for CIBP. Radioisotopes have a rapid onset of action, but data on NSCLC are limited and consist only of subgroup analyses . ZoledroImmune checkpoint inhibitors have become standard of care treatment for most patients with advanced NSCLC and result in durable responses in a subgroup of patients. For the subgroup of patients with oncogenic drivers, tyrosine kinase inhibitors often result in early and prolonged responses. For both classes of drugs, effects on CIBP have not been specifically reported. It is possible that in some patients, immune checkpoint inhibitors will not be very active in pain relief for CIBP, as Schmid et al. reported that efficacy of immunotherapy depends on the metastatic location: the treatment efficacy is less in bone lesions compared to lymph nodes . HoweverExperimental studies in animal models of CIBP have shown alterations in, e.g., astrocytes or in the sphingolipid metabolism in the spinal cord or showed the importance of connexins in the cell-cell communication with probable effects on CIBP. Recently, different studies focused on therapeutic options to block or alter these pathophysiological changes. Blockade of interleukine-6 signaling is promising as it could lead to prevention or delay of bone remodeling as well as decreased pain intensity .Some possible drawbacks for this systematic review exist. A point for discussion could be the chosen definition of early pain reduction (pain reduction within six weeks). We chose this upper limit to be as inclusive as possible to include treatment options that resulted in pain reduction within a relatively short term. Of note, for pain reduction treatment options within a shorter time frame , even less data is available. As discussed above, bisphosphonates have different activity on early CIBP reduction in breast- and prostate cancer compared with NSCLC, we did not broaden our inclusion criteria to include other tumor types –15, 41.Furthermore, as expected with over half of the included trials being single arm and/or not blinded and as shown by the Jadad score the methodological quality of most included trials is poor. Last, we did not include a formal test of heterogeneity, as only very heterogeneous trials met our inclusion criteria.In conclusion, despite the frequent occurrence of CIBP combined with the negative effects on QoL and OS, literature on the optimal treatment of CIBP in NSCLC is lacking. Most of the recommendations given in current guidelines are mainly based on data obtained in other tumors such as breast and prostate. Therefore, randomized trials evaluating treatment options with early pain relief for CIBP are necessary in lung cancer patients.No datasets were generated or analyzed for this study.Conceptualization: AB, BB, MB-E, LH. Formal Analysis: AB, BB. Writing—Original Draft Preparation: AB. Writing—Review and Editing: BB, MB-E, A-MD, LH. All authors agree to be accountable for the content of the work. All authors contributed to the article and approved the submitted version.A-MD has taken part on an advisory board for Roche.The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.