Nonparametric Two-sample Tests Research Articles

A two-sample nonparametric test for one-sided location-scale alternative

An increase in location is often accompanied by an increase in variability. Moreover, in randomized studies, the presence of heteroscedasticity can indicate a treatment effect. In these cases a location-scale test is appropriate. A common approach of a location-scale test is to use the sum of a location and a scale test statistic. In this study, we focus on one-sided location-scale tests. We introduce a one-sided Lepage-type test statistic. Since right-skewed data are common in many real-world applications, we design test statistics that are powerful also for right-skewed data. Moreover, this paper introduces both a maximum test and an adaptive test utilizing the one-sided Lepage-type test statistics. The limiting distributions of the maximum test statistics are also derived. We assess the performance of the different test statistics in various scenarios for continuous distributions through Monte Carlo simulations. Our simulation results demonstrate that the proposed new test statistics can significantly increase and stabilize statistical power, making it strong competitors to existing location-scale tests. We present practical illustrations based on example data and conclude with some final remarks.

Interference with activities of daily living according to pain level after breast surgery

BackgroundThe degree to which postoperative pain interferes with activities of daily living (ADLs) after breast surgery is unclear. We assessed the effect of pain on interference and analgesic use following lumpectomy and sentinel lymph node biopsy (lump-SLNB), and mastectomy. MethodsWe retrospectively identified consecutive patients who completed ≥1 post-discharge survey(s) on postoperative days (POD) 1–5 (lump-SLNB) and 1–10 (mastectomy) from 1/2019–12/2020. The highest pain score was used to categorize none/mild and moderate/severe pain groups. Interference was reported as “none/a little”, “somewhat”, and “quite a bit/very much”. Two-sample non-parametric tests compared baseline characteristics. Results1067 patients underwent lump-SLNB; 1219 underwent mastectomy—436 (41%) and 857 (70%) patients reported moderate/severe pain, of whom 190 (44%) and 121 (14%) rated interference as “none/a little”, 178 (41%) and 341 (40%) as “somewhat”, and 68 (16%) and 395 (46%) as “quite a bit/very much”, respectively. Patients with moderate/severe pain more frequently reported “quite a bit/very much” interference versus those with none/mild pain (lump-SLNB: 16% versus 1.1%, p<0.001; mastectomy: 46% versus 3.8%, p<0.001). Compared to POD1, the proportion of patients with the highest degree of interference declined on each subsequent POD in both surgical cohorts. Following mastectomy, median opioid use was 2 pills overall, and 6 pills for the highest interference group. ConclusionsInterference was more common in patients with moderate/severe pain; however, postmastectomy opioid use was low regardless of interference level. Our findings can inform expectations regarding postoperative pain, interference, and the feasibility of opioid-sparing recovery pathways. SynopsisPatients with moderate/severe pain after breast surgery experience more interference compared to those with none/mild pain. Postmastectomy opioid use is low, including patients reporting higher interference; most patients experience an early return to baseline function regardless of pain level.

Saddlepoint p-values for a class of location-scale tests

ABSTRACT The main idea of this paper is to approximate the exact p-value of a class of non-parametric, two-sample location-scale tests. In this paper, the most famous non-parametric two-sample location-scale tests are formulated in a class of linear rank tests. The permutation distribution of this class is derived from a random allocation design. This allows us to approximate the exact p-value of the non-parametric two-sample location-scale tests of the considered class using the saddlepoint approximation method. The proposed method shows high accuracy in approximating the exact p-value compared to the normal approximation method. Moreover, the proposed method only requires a few calculations and time, as in the case of the simulated method. The procedures of the proposed method are clarified through four sets of real data that represent applications for a number of different fields. In addition, a simulation study compares the proposed method with the traditional methods to approximate the exact p-value of the specified class of the non-parametric two-sample location-scale tests.

Impact of Neoadjuvant Chemoimmunotherapy on Surgical Outcomes and Time to Radiation in Triple-Negative Breast Cancer.

We examined the association between immunotherapy-containing and standard chemotherapy regimens with treatment delays and postoperative complications in stage II-III triple-negative breast cancer. The effect of immune-related adverse events (irAEs) was compared. We compared 139 women treated with neoadjuvant pembrolizumab plus chemotherapy (KEYNOTE-522 regimen) from August 2021 to September 2022 with 287 consecutive patients who received neoadjuvant chemotherapy alone prior to July 2021 and underwent surgery. Baseline characteristics, time to treatments, and surgical complications were compared using two-sample non-parametric tests. Linear regression evaluated association of irAEs with time to surgery and radiation. Logistic regression identified factors associated with surgical complications. Age, body mass index, race, American Society of Anesthesiologists (ASA) class, and mastectomy rates were similar among cohorts. No clinically relevant difference in time from end of neoadjuvant treatment to surgery was observed [KEYNOTE-522: median 32 (IQR 27, 43) days; non-KEYNOTE-522: median 31 (IQR 26, 37) days; P = 0.048]. Time to radiation did not differ (P = 0.7). A total of 26 patients (9%; non-KEYNOTE-522) versus 11 (8%; KEYNOTE-522) experienced postoperative complications (P = 0.6). In the KEYNOTE-522 cohort, 59 (43%) of 137 patients experienced 82 irAEs; 40 (68%) required treatment. Older age (P = 0.018) and ASA class 4 (P = 0.007) were associated with delays to surgery after adjusting for clinical factors. Experiencing ≥ 1 irAE was associated with delay to radiation (P = 0.029). IrAEs were not associated with surgical complications (P = 0.4). We observed no clinically meaningful difference between times to surgery/adjuvant radiation or postoperative complications and type of preoperative chemotherapy. IrAEs were associated with delay to adjuvant radiation but not with postoperative complications or delay to surgery.

Saddlepoint p-values for a class of location-scale tests under randomized block design

This paper deals with a class of nonparametric two-sample location-scale tests. The purpose of this paper is to approximate the exact p-value of the considered class under a randomized block design. The exact p-value of the considered class is approximated by the saddlepoint approximation method, also by the traditional method which is the normal approximation method. The saddlepoint approximation method is more accurate than the normal approximation method in approximating the exact p-value, and does not take a lot of time like the simulation method. This accuracy is proved by applying the mentioned methods to two real data sets and a simulation study.

Correcting a nonparametric two-sample graph hypothesis test for graphs with different numbers of vertices with applications to connectomics

Random graphs are statistical models that have many applications, ranging from neuroscience to social network analysis. Of particular interest in some applications is the problem of testing two random graphs for equality of generating distributions. Tang et al. (Bernoulli 23:1599–1630, 2017) propose a test for this setting. This test consists of embedding the graph into a low-dimensional space via the adjacency spectral embedding (ASE) and subsequently using a kernel two-sample test based on the maximum mean discrepancy. However, if the two graphs being compared have an unequal number of vertices, the test of Tang et al. (Bernoulli 23:1599–1630, 2017) may not be valid. We demonstrate the intuition behind this invalidity and propose a correction that makes any subsequent kernel- or distance-based test valid. Our method relies on sampling based on the asymptotic distribution for the ASE. We call these altered embeddings the corrected adjacency spectral embeddings (CASE). We also show that CASE remedies the exchangeability problem of the original test and demonstrate the validity and consistency of the test that uses CASE via a simulation study. Lastly, we apply our proposed test to the problem of determining equivalence of generating distributions in human connectomes extracted from diffusion magnetic resonance imaging at different scales.

An exact projection pursuit-based algorithm for multivariate two-sample nonparametric testing applicable to retrospective and group sequential studies

Nonparametric tests for equality of multivariate distributions are frequently desired in research. It is commonly required that test-procedures based on relatively small samples of vectors accurately control the corresponding Type I Error (TIE) rates. Often, in the multivariate testing, extensions of null-distribution-free univariate methods, e.g., Kolmogorov-Smirnov and Cramér-von Mises type schemes, are not exact, since their null distributions depend on underlying data distributions. The present paper extends the density-based empirical likelihood technique in order to nonparametrically approximate the most powerful test for the multivariate two-sample (MTS) problem, yielding an exact finite-sample test statistic. We rigorously apply one-to-one-mapping between the equality of vectors' distributions and the equality of distributions of relevant univariate linear projections. We establish a general algorithm that simplifies the use of projection pursuit, employing only a few of the infinitely many linear combinations of observed vectors' components. The displayed distribution-free strategy is employed in retrospective and group sequential manners. A novel MTS nonparametric procedure in the group sequential manner is proposed. The asymptotic consistency of the proposed technique is shown. Monte Carlo studies demonstrate that the proposed procedures exhibit extremely high and stable power characteristics across a variety of settings. Supplementary materials for this article are available online.

Feasibility and safety of planned early discharge following laparotomy in gynecologic oncology with enhanced recovery protocol including opioid-sparing anesthesia.

This study aims to evaluate the feasibility and safety of planned postoperative day 1 discharge (PPOD1) among patients who undergo laparotomy (XL) in the department of gynecology oncology utilizing a modified enhanced recovery after surgery (ERAS) protocol including opioid-sparing anesthesia (OSA) and defined discharge criteria. Patients undergoing XL and minimally invasive surgery (MIS) were enrolled in this prospective, observational cohort study after the departmental implementation of a modified ERAS protocol. The primary outcome was quality of life (QoL) using SF36, PROMIS GI, and ICIQ-FLUTS at baseline and 2- and 6-week postoperative visits. Statistical significance was assessed using the two-tailed Student's t-test and non-parametric Mann-Whitney two-sample test. Of the 141 subjects, no significant demographic differences were observed between the XL group and the MIS group. The majority of subjects, 84.7% (61), in the XL group had gynecologic malignancy [vs. MIS group; 21 (29.2%), p < 0.001]. All patients tolerated OSA. The XL group required higher intraoperative opioids [7.1 ± 9.2 morphine milligram equivalents (MME) vs. 3.9 ± 6.9 MME, p = 0.02] and longer surgical time (114.2 ± 41 min vs. 96.8 ± 32.1 min, p = 0.006). No significant difference was noted in the opioid requirements at the immediate postoperative phase and the rest of the postoperative day (POD) 0 or POD 1. In the XL group, 69 patients (73.6%) were successfully discharged home on POD1. There was no increase in the PROMIS score at 2 and 6 weeks compared to the preoperative phase. The readmission rates within 30 days after surgery (XL 4.2% vs. MIS 1.4%, p = 0.62), rates of surgical site infection (XL 0% vs. MIS 2.8%, p = 0.24), and mean number of post-discharge phone calls (0 vs. 0, p = 0.41) were comparable between the two groups. Although QoL scores were significantly lower than baseline in four of the nine QoL domains at 2 weeks post-laparotomy, all except physical health recovered by the 6-week time point. PPOD1 is a safe and feasible strategy for XL performed in the gynecologic oncology department. PPOD1 did not increase opioid requirements, readmission rates compared to MIS, and patient-reported constipation and nausea/vomiting compared to the preoperative phase.

Mutational Status is Associated with a Higher Rate of Pathologic Complete Response After Neoadjuvant Chemotherapy in Hormone Receptor-Positive Breast Cancer.

Pathologic complete response (pCR) to neoadjuvant chemotherapy (NAC) occurs in up to 20% of hormone receptor-positive (HR+)/human epidermal growth factor receptor 2-negative (HER2-) breast cancers. Whether this differs among BRCA mutation carriers is uncertain. This study compared pCR between BRCA1/2 mutation carriers and matched sporadic control subjects. From November 2013 to January 2022, this study identified 522 consecutive women with clinical stage I to III HR+/HER2- breast cancer treated with NAC and surgery. The study matched BRCA1/2 mutation carriers 1:2 to non-carriers in terms of age, clinical tumor (cT) and nodal (cN) stage, and differentiation. Two-sample non-parametric tests compared baseline characteristics. Multivariable logistic regression assessed pCR (i.e., ypT0/ispN0) by BRCA1/2 mutational status. Of the 522 women (median age, 50 years), 59 had BRCA1/2 mutations, 78% of which were clinically node positive. Anthracycline-based NAC was administered to 97%. More BRCA1/2 mutation carriers were younger, had cT1 tumors, and had poorly differentiated disease. After matching, 58 BRCA1/2 mutation carriers were similar to 116 non-carriers in terms of age (p = 0.6), cT (p = 0.9), cN stage (p = 0.7), and tumor differentiation (p > 0.9). Among the mutation carriers, the pCR rate was 15.5% for BRCA1/2, 38% (8/21) for BRCA1, and 2.7% (1/37) for BRCA2 versus 7.8% (9/116) for the non-carriers (p < 0.001). After NAC, 5 (41.7%) of the 12 BRCA1 mutation carriers converted to pN0 versus 10 (37%) of the 27 BRCA2 mutation carriers and 19 (20.9%) of the 91 non-carriers (p = 0.3). In the multivariable analysis, BRCA1 mutation status was associated with higher odds of pCR than non-carrier status (odds ratio [OR] 6.31; 95% confidence interval [CI] 1.95-20.5; p = 0.002), whereas BRCA2 mutation status was not (OR 0.45; 95% CI 0.02-2.67; p = 0.5). This study showed that BRCA1 mutation carriers with HR+/HER2- breast cancers have a higher rate of pCR than sporadic cancers and may derive greater benefit from chemotherapy. The use of NAC to downstage these patients should be considered.

Pathologic complete response after neoadjuvant systemic therapy for breast cancer in BRCA mutation carriers and noncarriers.

544 Background: Distinct pathological characteristics and mutational signatures in BRCA-associated breast cancers may result in differential response to chemotherapy but understanding of treatment response after neoadjuvant chemotherapy (NAC) is limited. We describe a single-institutional experience to compare rates of pathologic complete response (pCR) after NAC in BRCA carriers (g BRCA1/2) and noncarriers. Methods: From 11/2013 to 01/2022, 1426 consecutive women with clinical stage I-III breast cancer were treated with NAC followed by surgery. Baseline disease characteristics were compared between g BRCA1/2 and noncarriers using two-sample non-parametric tests. Conditional logistic regression was used to evaluate the association between BRCA status and pCR (i.e., ypT0/is pN0) adjusting for variables selected using backward elimination. Results: 92 (6.5%) and 73 (5.1%) had deleterious mutations in BRCA1 and BRCA2, respectively. Compared to noncarriers, g BRCA1/2 were younger (p&lt;0.001) with clinical T1 (p=0.002) and triple negative disease (TN) (57% vs 25% noncarriers, p&lt;0.001). Almost all patients received doxorubicin/cyclophosphamide/paclitaxel therapy (93%) with g BRCA1/2 more likely to receive carboplatin (p&lt;0.001). pCR rate was 42% of g BRCA1, 21% of g BRCA2, and 26% of noncarriers (p=0.001). Among clinically node positive (cN+) patients, nodal pCR was higher in g BRCA1/2 compared to noncarriers (53/96 (55%) vs 371/856 (43%), p=0.012). This difference was seen in ER+/HER2- (36% gBRCA1/2 vs 20% noncarriers; p=0.027) and TN subtypes (79% g BRCA1/2 vs 45% noncarriers; p&lt;0.001). Poorly differentiated tumors (p&lt;0.001), cN+ (p=0.010), lower cT stage (p=0.028), ductal histology (p&lt;0.001), TN and HER2+ receptor status (p&lt;0.001), absence of lymphovascular invasion (p&lt;0.001), carboplatin receipt (p=0.041), and BRCA1 status (p=0.001) were also associated with pCR on univariate analysis. After adjusting for differentiation and molecular subtype, BRCA1 status remained independently associated with pCR. Conclusions: g BRCA1 undergoing NAC have higher pCR rates relative to g BRCA2 and patients with sporadic disease. Across breast cancer subtypes, g BRCA1/2 more frequently converted to pN0. Defining biological mechanism for these findings would allow customization of treatment based on germline genetics. [Table: see text]

Impact of neoadjuvant chemo-immunotherapy on surgical outcomes and time to radiation in triple negative breast cancer.

555 Background: The impact of preoperative chemo-immunotherapy on surgical complications and timing of adjuvant therapy is not well understood. We examined the association between neoadjuvant pembrolizumab plus chemotherapy (NAC+P) compared to neoadjuvant chemotherapy alone (NAC) and treatment delays and postoperative complications in triple negative breast cancer (TNBC). The effect of immune-related adverse events (irAEs) on these outcomes was also compared. Methods: 143 women with stage II-III TNBC treated with NAC+P based on the KEYNOTE 522 (KN522) regimen from 8/2021-9/2022 were compared to 287 consecutive patients who received NAC prior to 7/2021. Baseline characteristics, time to treatments, and surgical complications were compared between KN522 and non-KN522, and among KN522 with/without irAEs using two-sample non-parametric tests. Linear regression evaluated association of irAEs with time to surgery and radiation. Conditional logistic regression identified factors associated with surgical complications among KN522 patients. Results: Age, BMI, race, ASA class, and mastectomy rates were similar among KN522 and non-KN522 patients. Interval from end of neoadjuvant treatment until surgery was one day longer for patients receiving NAC+P (median 32 days (IQR 27, 43) vs. 31 days (26, 37) non-KN522, p = 0.035). Time to radiation did not differ (p = 0.2). Postoperative complications occurred in 26 (9%) non-KN522 vs 12 (8.4%) of KN522 patients (p = 0.7) (Table). 55/144 (38%) of KN522 patients experienced 74 irAE; 42 (76%) required treatment. Pembrolizumab was held in 21 (38%) and discontinued permanently in 17 (31%). One KN522 patient experienced a fatal irAE (pneumonitis). Older age (coeff. 0.38, 95% CI 0.06-0.70, p = 0.02) and ASA class 4 (coeff. 53, 95% CI 14-92, p = 0.009) were associated with delays to surgery but irAE was not (p = 0.070). Neither presence of irAE (coeff. 6.8, 95% CI -0.74-14, p = 0.081) or number of irAEs (coeff. 4.1, 95% CI -1.2-9.3, p = 0.13) were associated with time to radiation. IrAEs were not associated with surgical complications (p = 0.5). Conclusions: Postoperative complications or delays to surgery or adjuvant radiation were similar in patients treated with NAC+P compared to NAC. IrAEs were not associated with delays to surgery or adjuvant radiation or postoperative complications. [Table: see text]

New nonparametric tests based on data depth for multivariate multi-sample location problem

Data depth based nonparametric tests for multivariate multisample location problem are provided which extend the depth based nonparametric test for multivariate two-sample location problem by Shirke and Khorate (2018) [Shirke, D.T. and Khorate, S.D., 2018. Two-sample nonparametric test for testing equality of locations based on data depth. Journal of the Indian Society for Probability and Statistics, 19, pp. 9-23.] to the multi-sample setting. Proposed tests are based on the correlations among the depths of data points. Power comparison of proposed tests with the existing depth based nonparametric tests is provided using a simulation study based on a permutation test. Simulation study reveals that the proposed tests give comparable powers compared to the existing data depth based nonparametric tests. Proposed tests are illustrated with real life data sets.

Abstract P425: Diets Enriched in Stearate, Palmitate, and Oleate Differentially Modulate Gut Microbiome Composition in Post-Menopausal Women

Objective: Dietary stearic acid (18:0), a major saturated fatty acid (SFA) in Western diets, does not raise plasma LDL-C concentrations relative to other shorter chain SFAs (palmitic [16:0], myristic [14:0] and lauric [12:0]), and is associated with lower CVD risk. We investigated whether the cardioprotective effects of 18:0 and its metabolic product oleic acid (18:1, a monounsaturated fatty acid) relative to 16:0 is mediated via alterations in the gut microbiome. Methods: Twenty mildly hypercholesterolemic (LDL-C&gt;100mg/dL) post-menopausal women (50-85 years; BMI 25-35kg/m 2 ) consumed each of 3 isocaloric diets for 5 weeks according to a randomized controlled cross-over design. Diets provided 55%E carbohydrate, 15%E protein and 30%E fat with half of the fat provided by 16:0, 18:0 or 18:1, respectively. Fecal samples were collected at the end of each diet phase and shotgun metagenomic sequencing was performed. Alpha diversity (Shannon index, Simpson index and observed counts) was compared between diet groups using two-sample non-parametric tests. Beta-diversity was assessed using Bray-Curtis dissimilarity with PERMANOVA test. Differences in metagenomic features by diet group were evaluated using Partial Least Squares-Discriminant Analysis (PLS-DA) models of pairwise diet differences developed using the R package ‘mixOmics’. Results: No significant differences were observed in alpha and beta diversity between diet groups. However, the variability in microbial relative abundance was greater after participants consumed the 18:0 and 18:1 diets compared to the 16:0 diet. Specifically, mean relative abundance of Bacterioides dorei , Bacteroides vulgatus , and Ruminococcus were highest after participants consumed the 18:0 relative to the 16:0 diet and intermediate after the 18:1 diet. The relative abundance of Coprococcus , Parabacteroides distasonis , Coprobacillus and Blautia were higher after participants consumed the 18:1 relative to the 16:0 diet. These microbial species have been associated with several anti-inflammatory pathways and decreased bacterial lipopolysaccharide production. Conclusion: These data document modest but distinctive effects of dietary SFAs on gut microbiome composition and functionality, suggesting that the favorable effects of 18:0 and 18:1 relative to 16:0 on CVD risk factors could be mediated, in part, by the gut microbiome.

Two-sample nonparametric test for proportional reversed hazards

In the last few decades, several works have been undertaken in the context of proportional reversed hazard rates (PRHR) but any specific statistical methodology for the PRHR hypothesis is absent in the literature. This paper proposes a two-sample nonparametric test based on two independent samples to verify the PRHR assumption. Based on a consistent U-statistic three statistical methodologies have been developed exploiting U-statistics theory, jackknife empirical likelihood and adjusted jackknife empirical likelihood method. A simulation study has been performed to assess the merit of the proposed test procedure. Finally, the proposed procedure is applied to a data set in the context of brain injury-related biomarkers and a data set related to Duchenne muscular dystrophy.

AUGUST: An Interpretable, Resolution-based Two-sample Test

Two-sample testing is a fundamental problem in statistics. While many powerful nonparametric methods exist for both the univariate and multivariate context, it is comparatively less common to see a framework for determining which data features lead to rejection of the null. In this paper, we propose a new nonparametric two-sample test named AUGUST, which incorporates a framework for interpretation while maintaining power comparable to existing methods. AUGUST tests for inequality in distribution up to a predetermined resolution using symmetry statistics from binary expansion. Designed for univariate and low to moderate-dimensional multivariate data, this construction allows us to understand distributional differences as a combination of fundamental orthogonal signals. Asymptotic theory for the test statistic facilitates p-value computation and power analysis, and an efficient algorithm enables computation on large data sets. In empirical studies, we show that our test has power comparable to that of popular existing methods, as well as greater power in some circumstances. We illustrate the interpretability of our method using NBA shooting data.

Nonparametric Two-Sample Tests of High Dimensional Mean Vectors via Random Integration

Testing the equality of the means in two samples is a fundamental statistical inferential problem. Most of the existing methods are based on the sum-of-squares or supremum statistics. They are possibly powerful in some situations, but not in others, and they do not work in a unified way. Using random integration of the difference, we develop a framework that includes and extends many existing methods, especially in high-dimensional settings, without restricting the same covariance matrices or sparsity. Under a general multivariate model, we can derive the asymptotic properties of the proposed test statistic without specifying a relationship between the data dimension and sample size explicitly. Specifically, the new framework allows us to better understand the test’s properties and select a powerful procedure accordingly. For example, we prove that our proposed test can achieve the power of 1 when nonzero signals in the true mean differences are weakly dense with nearly the same sign. In addition, we delineate the conditions under which the asymptotic relative Pitman efficiency of our proposed test to its competitor is greater than or equal to 1. Extensive numerical studies and a real data example demonstrate the potential of our proposed test. Supplementary materials for this article are available online.

Bi-Invariant Dissimilarity Measures for Sample Distributions in Lie Groups

Data sets sampled in Lie groups are widespread, and as with multivariate data, it is important for many applications to assess the differences between the sets in terms of their distributions. Indices for this task are usually derived by considering the Lie group as a Riemannian manifold. Then, however, compatibility with the group operation is guaranteed only if a bi-invariant metric exists, which is not the case for most noncompact and noncommutative groups. We show here that if one considers an affine connection structure instead, one obtains bi-invariant generalizations of well-known dissimilarity measures: Hotelling statistic, Bhattacharyya distance, and Hellinger distance. Each of the dissimilarity measures matches its multivariate counterpart for Euclidean data and is translation invariant, so that biases, e.g., through an arbitrary choice of reference, are avoided. We further derive nonparametric two-sample tests that are bi-invariant and consistent. We demonstrate the potential of these dissimilarity measures by performing group tests on data of knee configurations and epidemiological shape data. Significant differences are revealed in both cases.

A Nonparametric Two-Sample Test for Scale Using Linear Combination of Quadratic and Linear Function of Placements

A Nonparametric Two-Sample Test for Scale Using Linear Combination of Quadratic and Linear Function of Placements

Brain proteome profiling implicates the complement and coagulation cascade in multiple system atrophy brain pathology.

BackgroundMultiple system atrophy (MSA) is a rare, progressive, neurodegenerative disorder presenting glia pathology. Still, disease etiology and pathophysiology are unknown, but neuro-inflammation and vascular disruption may be contributing factors to the disease progression. Here, we performed an ex vivo deep proteome profiling of the prefrontal cortex of MSA patients to reveal disease-relevant molecular neuropathological processes. Observations were validated in plasma and cerebrospinal fluid (CSF) of novel cross-sectional patient cohorts.MethodsBrains from 45 MSA patients and 30 normal controls (CTRLs) were included. Brain samples were homogenized and trypsinized for peptide formation and analyzed by high-performance liquid chromatography tandem mass spectrometry (LC–MS/MS). Results were supplemented by western blotting, immuno-capture, tissue clearing and 3D imaging, immunohistochemistry and immunofluorescence. Subsequent measurements of glial fibrillary acid protein (GFAP) and neuro-filament light chain (NFL) levels were performed by immunoblotting in plasma of 20 MSA patients and 20 CTRLs. Finally, we performed a proteome profiling of 144 CSF samples from MSA and CTRLs, as well as other parkinsonian disorders. Data were analyzed using relevant parametric and non-parametric two-sample tests or linear regression tests followed by post hoc tests corrected for multiple testing. Additionally, high-throughput bioinformatic analyses were applied.ResultsWe quantified more than 4,000 proteins across samples and identified 49 differentially expressed proteins with significantly different abundances in MSA patients compared with CTRLs. Pathway analyses showed enrichment of processes related to fibrinolysis and complement cascade activation. Increased fibrinogen subunit β (FGB) protein levels were further verified, and we identified an enriched recognition of FGB by IgGs as well as intra-parenchymal accumulation around blood vessels. We corroborated blood–brain barrier leakage by a significant increase in GFAP and NFL plasma levels in MSA patients that correlated to disease severity and/or duration. Proteome profiling of CSF samples acquired during the disease course, confirmed increased total fibrinogen levels and immune-related components in the soluble fraction of MSA patients. This was also true for the other atypical parkinsonian disorders, dementia with Lewy bodies and progressive supra-nuclear palsy, but not for Parkinson’s disease patients.ConclusionOur results implicate activation of the fibrinolytic cascade and immune system in the brain as contributing factors in MSA associated with a more severe disease course.Supplementary InformationThe online version contains supplementary material available at 10.1007/s00018-022-04378-z.

Characterizing Outcomes of Medial and Lateral Perforators in Deep Inferior Epigastric Perforator Flaps.

Perforators are typically found in rows in the deep inferior epigastric perforator (DIEP) flap. As methods to assess flap perfusion continue to improve, surgeons may be more likely to select perforators traditionally avoided. The purpose of this article is to describe clinical outcomes based on row and number of perforators to reevaluate flap and abdominal donor site morbidity. A retrospective analysis was performed on patients who underwent breast reconstruction with DIEP flaps by four microsurgeons from 2013 to 2020. The row and number of perforators were determined from operative reports. Chi-square and t-test or nonparametric Fisher's exact test and Wilcoxon two-sample test were used for discrete and continuous variable, respectively, as applicable. Logistic regression was used for multivariable analyses. Of 628 flaps, 305 were medial row (58.7%), 159 were lateral row (30.6%), and 55 had both rows (10.6%). Partial flap loss was higher in both rows (p = 0.003). Fat necrosis was higher with medial (p = 0.03) and both rows (p = 0.01) when compared with lateral using multivariable analysis. Hernia or bulge was higher in lateral row flaps (lateral: 8/157, 5.1%; medial, 5/299, 1.7%; both, 0/55; p = 0.05); however, mesh was more commonly used in both row flaps (p = 0.05). There was no difference in fat necrosis or abdominal morbidity between single and multiple perforators. There was no difference in fat necrosis based on the number or row of perforators. The lateral row provides adequate perfusion but may be associated with an elevated risk of hernia or bulge. Patients may benefit from mesh, especially when both rows are dissected.