Weight Regain After Lifetype Interventions is Associated with Higher

Introduction

Obesity is a serious threat issue for a large spectrum of persistent ailments, together with sort 2 diabetes, heart problems, and metabolic dysfunction–related steatotic liver illness (MASLD), and its international prevalence continues to rise.¹ MASLD, not too long ago redefined to emphasise its shut hyperlink with metabolic dysfunction, has change into essentially the most prevalent persistent liver illness globally, affecting roughly 30% of the grownup inhabitants.^2–4 Beyond progressive liver damage—starting from steatosis to steatohepatitis, fibrosis, cirrhosis, and hepatocellular carcinoma—MASLD can also be strongly related to extrahepatic issues, notably heart problems,⁵ sort 2 diabetes,⁶ stroke,⁷ and persistent kidney illness.⁸

MASLD represents hepatic manifestation of systemic metabolic dysfunction and is intently intertwined with weight problems, insulin resistance, dyslipidemia, and persistent low-grade irritation.⁹ Sustained weight administration is due to this fact a cornerstone of each obesity- and MASLD-related illness prevention and therapy. Considerable weight reduction may be achieved by means of numerous approaches akin to way of life modifications,^10,11 pharmaceutical interventions,^12,13 or bariatric surgical process.^14,15 A weight lack of 5% has proved to be a benchmark of enhancements in well being outcomes, together with cardiovascular advantages¹⁶ and liver steatosis enhancements,¹⁷ making it at present the primary purpose in weight administration interventions.¹⁸ However, weight reduction by way of completely different methods is usually adopted by a subsequent weight regain amongst most people,¹⁹ which poses a posh problem for long-term weight administration.

The metabolic penalties of weight regain stay incompletely understood. While some research, together with reanalysis of the Look AHEAD trial, recommend that the advantages of weight reduction persist regardless of subsequent regain,^20,21 others have linked weight regain to elevated mortality and cardiovascular threat.^22,23 Importantly, present proof is basically derived from cardiovascular or diabetes-focused outcomes, with restricted consideration to liver-related endpoints or validated noninvasive indices of MASLD severity. Moreover, weight problems is characterised by persistent low-grade irritation, elevating concern that weight regain might reactivate inflammatory pathways and speed up metabolic deterioration, doubtlessly exacerbating insulin resistance and MASLD development.²⁴ Whether weight regain finally confers internet metabolic hurt in contrast with weight stability in people with weight problems due to this fact stays an unresolved scientific query.

Notably, knowledge are notably scarce concerning the long-term metabolic and hepatic impression of weight regain following lifestyle-induced weight reduction, regardless of way of life intervention being essentially the most broadly advisable first-line remedy. This lack of readability represents a crucial hole in data with direct implications for affected person counseling, long-term administration methods, and threat stratification in weight problems and MASLD. To handle this hole, the current examine investigates whether or not weight regain after reaching ≥5% weight reduction by means of way of life interventions is related to hostile metabolic outcomes, in contrast with sustained weight stability over a three-year follow-up interval. By comprehensively evaluating metabolic parameters and noninvasive indices associated to MASLD, this examine goals to make clear the scientific significance of weight regain and inform long-term weight administration methods in people with weight problems.

Methods

Study Population

The retrospective examine included sufferers exhibiting both weight regain or sustained weight over a steady interval of three years earlier than their preliminary analysis of weight problems (BMI ≥ 30 kg/m²). These sufferers have been sourced from the specialised division of weight problems of Ruijin Hospital, affiliated to Shanghai Jiao Tong University School of Medicine, from September 2018 to September 2023. All contributors have been Han Chinese aged 18–45 years. In this examine, “weight regain” was outlined as a documented historical past of dropping at the least 5% of their preliminary weight by the use of way of life intervention (together with cognitive restraint and train) adopted by a return to or exceeding their preliminary weight, and “weight sustain” was outlined as experiencing weight fluctuations inside a variety of 5% (steady-obese state of sufferers). The exclusion standards have been as follows: 1) weight problems secondary to recognized genetic syndromes (for instance, Prader–Willi syndrome), endocrine problems (for instance, Cushing’s syndrome), or medicine use (for instance, corticosteroids, antidepressants); 2) use of any pharmacologic therapy inside three months previous the examine go to; 3) documented historical past of alcohol, tobacco, or substance abuse; 4) body-weight trajectories inconsistent with the above definitions; or 5) people with out dependable weight trajectory historical past. Conduction of the examine together with the waiver of knowledgeable consent have been accepted by the Institutional Review Board of the Ruijin Hospital, Shanghai Jiao Tong University School of Medicine (Approval quantity: Ethics 2023[411]).

Sample Size

Sample measurement was estimated primarily based on preliminary findings from the Genetics of Obesity in Chinese Youngs (GOCY) cohort (ClinicalTrials.gov identifier: NCT01084967). The imply ALT concentrations have been roughly 65 ± 45 IU/L amongst people with weight regain and 45 ± 45 IU/L amongst these with sustained weight problems. Assuming α = 0.05, energy = 0.80, and a 2:1 allocation ratio, the minimal required pattern sizes have been calculated as 120 and 60 for the 2 teams, respectively (complete = 180).

Measurements

Clinical info was retrieved from digital medical information. Height and physique weight have been measured with contributors sporting mild clothes and no sneakers, recorded to the closest 0.1 cm and 0.1 kg, respectively. BMI (kg/m²) was calculated as physique weight divided by the sq. of peak.All contributors underwent an oral glucose tolerance take a look at (OGTT). Fasting and 2-hour glucose values have been decided utilizing an automatic analyzer (AU5800; Beckman Coulter, CA, USA). The similar system measured liver enzymes (alanine aminotransferase [ALT], aspartate aminotransferase [AST], alkaline phosphatase [AKP], and γ-glutamyl transpeptidase [GGT]), creatinine, uric acid, and serum lipids (triglycerides [TG], complete ldl cholesterol [TC], high-density ldl cholesterol [HDL-C] and low-density lipoprotein ldl cholesterol [LDL-C]). Fasting and 2-hour insulin concentrations have been measured by double-antibody radioimmunoassay (DSL, Webster, TX, USA). Hemoglobin A1c (HbA1c) was quantified by high-performance liquid chromatography utilizing the VARIANT II analyzer (Bio-Rad Laboratories). Thyroid hormones—free triiodothyronine (fT3), free thyroxine (fT4), and thyroid-stimulating hormone (TSH)—have been decided utilizing chemiluminescent immunoassay (Architect i2000sr; Abbott Laboratories, IL, USA).

Assessment of Insulin Resistance, Insulin Sensitivity, and β-Cell Function

Insulin resistance and insulin sensitivity have been assessed utilizing each fasting- and OGTT-derived indices. Homeostasis mannequin evaluation of insulin resistance (HOMA-IR) was calculated as fasting plasma glucose (mmol/L) × fasting insulin (μU/mL) / 22.5. Homeostasis mannequin evaluation of β-cell operate (HOMA-β) was calculated as 20 × fasting insulin (μU/mL) / [fasting plasma glucose (mmol/L) − 3.5]. Whole-body insulin sensitivity was estimated utilizing the Matsuda insulin sensitivity index (ISI), additionally known as ISI (0, 120), calculated utilizing the Matsuda formulation primarily based on glucose and insulin values obtained throughout OGTT.²⁵ The disposition index (DI) was calculated because the ratio of HOMA-β to HOMA-IR and was used to evaluate β-cell operate adjusted for insulin resistance, reflecting the flexibility of pancreatic β-cells to compensate for insulin resistance.

Non-Invasive Tests Evaluating Liver Steatosis and Liver Fibrosis

The Dallas Steatosis Index (DSI),²⁶ Nonalcoholic Fatty Liver Disease Fibrosis Score (NFS) and Fibrosis-4 (FIB-4) index have been calculated in accordance with validated formulation (NFS and FIB-4 formulation see Supplementary Table 1). Established cutoff values have been utilized to establish superior fibrosis, with NFS ≥ −1.45 and FIB-4 ≥ 1.3, in accordance with present scientific pointers.²⁷

Acquisition and Preservation of Liver Specimens

Liver biopsy samples have been obtained from contributors who additionally took half within the scientific trial Efficacy and Mechanism Study of Bariatric Surgery to Treat Moderate to Severe Obesity in Han Chinese Population (ClinicalTrials.gov identifier: NCT02653430). Written knowledgeable consent was obtained from all donors. Biopsy specimens collected throughout laparoscopic sleeve gastrectomy have been instantly snap-frozen in liquid nitrogen and saved till RNA isolation.

Liver Histology and NAFLD Activity Score (NAS)

Liver biopsy specimens have been evaluated by two blinded hepatopathologists. Steatosis (0–3), lobular irritation (0–3), and hepatocellular ballooning (0–2) have been scored to calculate the NAS, with fibrosis assessed individually (0–4) in accordance with the NASH Clinical Research Network standards. Discrepancies have been resolved by consensus.

RNA Extraction and cDNA Preparation

Total RNA was extracted from liver tissue utilizing the Eastep Super Total RNA Extraction Kit (Promega, ls1040). Complementary DNA (cDNA) was synthesized from 1 µg of complete RNA utilizing the HiScript III All-in-One RT SuperMix (Vazyme, China) in accordance with the producer’s directions.

Bulk-RNA Sequencing and Analysis

RNA integrity was verified utilizing an Agilent 2100 Bioanalyzer (Agilent Technologies, CA, USA) and agarose gel electrophoresis. Poly(A) mRNA was remoted from complete RNA utilizing Oligo(dT) magnetic beads and fragmented into brief items previous to reverse transcription with the NEBNext Ultra RNA Library Prep Kit for Illumina (NEB #7530). The ensuing cDNA fragments underwent end-repair, A-tailing, and adaptor ligation, adopted by purification with AMPure XP beads and PCR amplification. Sequencing was carried out on an Illumina NovaSeq 6000 platform (Gene Denovo Biotechnology, Guangzhou, China). Principal element evaluation (PCA) was carried out utilizing the R package deal gmodels. Differentially expressed genes have been recognized with DESeq2 utilizing a false discovery price (FDR) < 0.05 and absolute fold change ≥ 2. Gene set enrichment evaluation (GSEA) was carried out with MSigDB to find out considerably enriched Gene Ontology (GO) phrases.

Real-Time PCR

Real-time PCR was carried out on an ABI system (Life Technologies) utilizing ChamQ Universal SYBR qPCR Master Mix (Vazyme, China). Reactions have been run in duplicate in 384-well plates. Relative expression ranges have been calculated utilizing the two^−ΔΔCt methodology after normalization to the GAPDH housekeeping gene. Primer sequences are listed in Supplementary Table 2.

Immunohistochemistry (IHC) Staining

Immunohistochemical staining was carried out with major monoclonal antibodies concentrating on CD11b (ab133357, Abcam, UK), CD68 (76437S, Cell Signaling Technology, USA), and IL-6 (ab233706, Abcam, UK) and Anti-alpha clean muscle Actin (ab7817, Abcam, UK). Formalin-fixed, paraffin-embedded liver sections have been dewaxed, subjected to antigen retrieval, and blocked for 1 h in phosphate-buffered saline containing 5% bovine serum albumin (BSA; Sigma, USA). Primary antibodies have been incubated in a single day at 4 °C, adopted by washing and incubation with HRP-conjugated secondary antibodies for 30 min at room temperature. Visualization was carried out utilizing the REAL™ EnVision™ system (DAKO, Denmark). Slides have been examined underneath a lightweight microscope (TissueFAXS viewer). The built-in optical densities (IODs) of goal proteins calculated by ImageJ software program.

Statistical Analysis

Continuous variables with regular distribution are expressed as imply ± normal deviation; non-normally distributed knowledge are offered as median (interquartile vary). Between-group comparisons have been carried out utilizing the t-test or the Mann–Whitney U-test, as applicable. Categorical knowledge have been expressed as counts (percentages) and in contrast utilizing the chi-square take a look at.

To decrease confounding from demographic elements, propensity rating matching (PSM) was carried out utilizing nearest-neighbor matching with out alternative, adjusting for age, intercourse, physique weight, and BMI. An absolute standardized imply distinction (SMD) < 0.2 was thought-about indicative of ample steadiness. Subgroup analyses have been additional carried out in accordance with intercourse.

Data evaluation was carried out with R model 4.2.0 and GraphPad Prism 9 (GraphPad Software Inc., San Diego, CA, USA). Propensity rating matching was carried out with the R package deal “Matching”.²⁸ A two-sided P worth of lower than 0.05 was thought to be being statistically vital.

Results

Recruitment

A complete of 1068 sufferers aged 18–45 years have been initially identified with weight problems on the specialised division of weight problems of Ruijin Hospital, affiliated with Shanghai Jiao Tong University School of Medicine, from September 2018 to September 2023, a complete of 213 sufferers have been included on this examine. Figure 1 confirmed the circulate diagram of the examine protocol.

Figure 1 Patient flow chart.

Basic Characteristics and Clinical Assessments of All Participants

A total of 157 patients with weight regain and 56 patients with weight sustain were included in this study. Patients in the weight regain group were younger [mean age, 27.32 (5.28) vs 29.82 (6.58), P=0.006]. No significant differences were observed between the two groups in metabolic and endocrine parameters, including glycemic indices, insulin resistance and sensitivity, lipid profiles, and thyroid function (Table 1). In addition, liver and renal operate parameters in addition to inflammatory markers have been comparable between the 2 teams (Table 2).

Table 1 Metabolic and Endocrine Parameters of Weight Regain and Sustain Group, Raw and Propensity Score-Matched Data

Table 2 Organ Function and Inflammatory Parameters of Weight Regain and Sustain Group, Raw and Propensity Score-Matched Data

After PSM of age, sex, weight, and BMI, 50 cases were matched each group. SMDs of the matched variables were all below 0.2, indicating a balanced match. Among all the matched patients, the weight regain group showed significantly higher ALT levels [median (IQR), 59.00 (40.25, 94.00) IU/L vs 41.00 (29.50, 58.00) IU/L, P=0.007] and higher AST levels [median (IQR), 33.50 (24.25, 55.00) IU/L vs 26.00 (21.00, 35.75) IU/L, P=0.041] (Table 2). No statistically vital distinction was demonstrated between the 2 teams by way of different biochemical indexes.

Subgroup Analysis of Clinical Characteristics

We additional analyzed the traits categorized by intercourse. Male sufferers (Table 3) confirmed greater weight at baseline within the weight regain group [mean weight (SD), 125.83 (17.26) kg vs 116.40 (19.89) kg, P=0.015], whereas feminine sufferers (Table 4) have been youthful within the weight regain group [mean age (SD), 27.31 (5.06) vs 30.38 (7.45), P=0.025]. Also, male sufferers within the weight regain group confirmed greater degree of ALT [median (IQR), 64.00 (42.00, 109.00) IU/L vs 47.00 (35.00, 65.00) IU/L, P=0.017]. The two teams have been comparable with respect to different biochemical indices in each female and male sufferers earlier than matching (Supplementary Tables 3 and 4).

Table 3 Subgroup Analysis of Organ Function Parameters of Male Patients, Raw and Propensity Score-Matched Data

Table 4 Subgroup Analysis of Organ Function Parameters of Female Patients, Raw and Propensity Score-Matched Data

After PSM, 31 male patients from each group were matched (Table 3). In the subgroup evaluation, a stronger pattern was noticed within the measurement of ALT degree [median (IQR), 88.00 (57.00, 137.50) IU/L vs 47.00 (35.00, 65.00) IU/L, P<0.001] and AST degree [median (IQR), 46.00 (31.50, 60.00) IU/L vs 30.00 (23.00, 36.50) IU/L, P=0.004] in male sufferers. In feminine sufferers, 30 sufferers from the load regain group and 15 sufferers from the load maintain group have been matched (Table 4). Unlike the male sufferers, feminine sufferers with weight regain confirmed greater uric acid degree [mean (SD), 429.70 (74.05) μmol/L vs 380.87 (66.15) μmol/L, P=0.037] than these with sustained weight. Other metabolic parameters didn’t differ considerably between the 2 teams in each sexes after matching (Supplementary Tables 3 and 4).

Non-Invasive Scoring Systems (DSI, NFS, FIB-4) Evaluating Hepatic Dysfunction

Since ALT and AST ranges differed between the 2 teams, we additionally calculated DSI, NFS and FIB-4 scores to additional consider hepatic dysfunction related to steatosis or fibrosis.

DSI is a validated device for the correct detection of early-grade hepatic steatosis.²⁶ In the general inhabitants, DSI values didn’t differ considerably between the 2 teams, earlier than or after PSM (Table 2). In male sufferers, greater DSI was present in weight regain group [median (IQR), 0.20 [−0.38, 0.72] vs −0.19 [−0.88, 0.34], P=0.043] (Table 3). However, this distinction was now not noticed after PSM. Also, DSI was discovered comparable between teams in feminine sufferers, earlier than and after matching (Table 4).

NFS and FIB-4 are basic non-invasive scoring methods evaluating liver fibrosis. Among these with out there knowledge, 23.5% within the weight regain group and 27.5% within the weight maintain group had NFS values above the cutoff. Meanwhile, 4.4% within the weight regain group and three.9% within the weight maintain group had FIB-4 values above the cutoff. There have been no vital variations within the optimistic charges between the teams for both scoring system (Supplementary Table 5). Additionally, no variations have been noticed between the teams after matching (Supplementary Table 6).

In male sufferers, amongst these with knowledge, 26.3% within the weight regain group and 29.0% within the weight maintain group had NFS values above the cutoff. 8.5% within the weight regain group had FIB-4 values above the cutoff, in comparison with 6.5% within the weight maintain group. No vital variations have been discovered between the teams, earlier than or after matching (Supplementary Tables 7 and 8).

Sensitivity Analysis Assessing the Impact of Excess Weight Gain

Notably, the method of weight acquire itself might contribute to hepatic alterations.²⁹ In our examine, some people within the weight regain group skilled weight acquire that exceeded their baseline ranges, which can have contributed to the noticed liver dysfunction. Hence, we carried out a one-way sensitivity evaluation to evaluate the impression of the surplus weight acquire on the reliability of our findings.

We excluded sufferers whose weight exceeded greater than 5% of baseline after weight regain. After the adjustment, a complete of 106 sufferers (54.7% feminine) have been included within the weight regain group. ALT and AST ranges weren’t considerably greater within the weight regain group, earlier than or after PSM, within the general pattern. However, ALT degree remained considerably greater within the male sufferers of weight regain group [median (IQR), 60.00 (41.00, 126.00) IU/L vs 43.00 (32.50, 54.50) IU/L, P=0.015] after PSM (weight regain: weight maintain= 27:27) (Table 5), whereas the degrees of liver enzymes remained equal in feminine sufferers of each teams, earlier than and after matching (Supplementary Table 9).

Table 5 Clinical Features of Weight Regain and Sustain Group in Sensitivity Analysis

Correlation of Liver Dysfunction and Inflammatory Status in Weight Regain Group

To gain a deeper understanding of the relationship between weight regain and liver metabolism, we conducted further analyses in a subset of bariatric surgery patients who underwent liver biopsy. A total of 19 male patients (weight regain: weight sustain=13:6) and 23 female patients (weight regain: weight sustain=20:3) had undergone bariatric surgery and provided liver biopsy samples. We matched liver specimen providers’ basic characteristics to avoid confounding factors. First, we performed histopathological assessment using NAS on the matched biopsy samples. Scores for steatosis, lobular inflammation, ballooning, and fibrosis stage were comparable between groups in male patients (individual data for all participants, including a limited number of female samples, are provided in Supplementary Table 10), indicating no major disparity in structural liver injury at the time of biopsy.

Given the absence of pronounced histopathological divergence in this analyzable cohort yet persistent clinical biochemical differences, we performed RNA-seq analysis on liver samples of matched male patients to identify earlier, pathway-level alterations that might explain the observed phenotype (Figure 2A). PCA was utilized to visualise the pattern distribution patterns (Figure 2B). Among the genes that have been recognized, 296 have been upregulated and 201 have been downregulated within the weight regain group (Figure 2C). Furthermore, evaluation of the gene transcription profiles by means of Gene Ontology revealed that the differential genes within the two teams have been primarily enriched in immune-related pathways (Figure 2D). GSEA evaluation of various genes indicated that a lot of the upregulated pathways within the weight regain group have been related to macrophage activation in addition to monocyte chemotaxis (Figure 2E and F). We subsequently verified that the expression of immune-related gene CD11B (P=0.016) was up-regulated in liver tissues of the load regain group by mRNA quantification; whereas expression of CD68 tended to be greater within the weight regain group (P=0.0536) (Figure 2G). Additionally, the mRNA expression of α-SMA, a marker for hepatic stellate cell activation and fibrosis, confirmed no vital distinction between the 2 teams. Immunohistochemical staining was carried out to detect the expression of CD11B, CD68, IL-6 and α-SMA in liver tissues (Figure 2H). As proven, the expression of CD11B, CD68 and IL-6 was greater within the weight regain group in contrast with the load maintain group. Consistently, immunohistochemical evaluation of α-SMA additionally revealed no apparent distinction in staining between the teams (Figure 2I).

Figure 2 Correlation of liver dysfunction and inflammatory status in male patients of weight regain group. (A) Flow chart of the experiment. (B) Principal component analysis (PCA) of the transcriptome profiles between weight sustain and weight regain group. (C) Volcano plot of differentially expressed genes between weight sustain and weight regain group. (D) The top10 regulated GO pathways between weight sustain and weight regain group. (E) and (F) GSEA results of microphage activation (E) and monocyte chemotaxis (F). (G) The mRNA expression of pro-inflammatory cytokines and fibrosis related genes between male subgroups. (H) Immunohistochemistry (IHC) staining representative images of CD11B, CD68, IL6 and α-SMA in male liver tissues (scale bar = 100 μm). (I) Quantitative analysis of images from (H). n=4. Data are shown as mean ± SEM. Statistical significance was assessed by unpaired two-sided Student’s t-test.

Discussion

Weight loss is actively pursued by a substantial proportion of the global population, with an estimated 42% of adults attempting to lose weight each year.³⁰ However, long-term weight upkeep stays difficult, and most people finally expertise partial or full weight regain.³¹ While sustained weight lack of ≥5% by means of way of life interventions improves metabolic well being and high quality of life,³² the organic penalties of subsequent weight regain stay incompletely understood.

In our present pilot examine, after adjustment for confounders, people with weight regain exhibited comparable glycemic indices, insulin resistance, lipid profiles, and endocrine parameters relative to these with sustained weight standing. In distinction, hepatic transaminase ranges have been modestly however constantly greater within the weight regain group. These findings recommend that, within the brief time period, weight regain might not result in broad deterioration of classical metabolic biomarkers however might mirror early liver-related biochemical alterations.

Elevated liver enzymes are generally interpreted as indicators of hepatocellular stress or damage. Although these modifications don’t essentially point out overt liver illness, they could mirror subclinical hepatic vulnerability. Our findings lengthen observations from animal fashions,^33–35 during which weight biking has been related to hepatic irritation and damage, to a human cohort. Importantly, this affiliation was noticed within the absence of great variations in glucose homeostasis or lipid metabolism, highlighting the liver as a doubtlessly delicate organ within the context of weight regain.

Sex-stratified analyses after propensity rating matching revealed distinct patterns: males with weight regain confirmed greater hepatic enzyme ranges, whereas ladies exhibited greater serum uric acid concentrations. These variations recommend sex-specific biochemical responses, although underlying mechanisms can’t be decided from the present dataset. Also, sex-stratified findings are reported descriptively and must be interpreted cautiously given restricted feminine pattern measurement.

At the tissue degree, people with weight regain exhibited elevated hepatic expression of macrophage markers CD68 and CD11B, whereas established indices of steatosis and fibrosis—together with NAS rating, Dallas Steatosis Index, NFS, FIB‑4 index, and α‑SMA—didn’t differ considerably. This sample is per early hepatic alterations characterised by innate immune activation, previous detectable steatosis or fibrotic reworking. Innate immunity performs a central function in liver irritation, with Kupffer cells and monocyte-derived macrophages contributing to pro-inflammatory signaling and immune recruitment.^36,37 Experimental research present that macrophage activation markers akin to CD68 and CD11b enhance throughout early diet-induced liver damage earlier than vital fibrosis develops,³⁸ and human research equally affiliate macrophage activation with hepatic damage metrics in MASLD.³⁹ Thus, the selective upregulation of macrophage-related genes noticed within the weight regain group is suitable with an early inflammatory response, per fashions proposing that immune activation precedes structural pathology.⁴⁰

Emerging proof on innate immune reminiscence in metabolic tissues gives further context. Prior publicity to obesity-associated inflammatory stimuli has been proposed to induce sturdy practical reprogramming of innate immune cells, leading to heightened responsiveness upon renewed metabolic problem.⁴¹ In adipose tissue, such imprinting persists regardless of weight reduction and may be reactivated upon renewed problem.^42,43 In the current examine, elevated hepatic macrophage marker expression, alongside tendencies towards greater IL-6 and IL-1B and decrease IL-10, is suitable with—however doesn’t show—the speculation that prior weight problems primes hepatic innate immune cells. This interpretation must be thought-about hypothesis-generating, with longitudinal research wanted to make clear whether or not immune reminiscence contributes to liver vulnerability throughout weight regain.

The present examine was designed as a pilot investigation to discover the metabolic and hepatic penalties of weight regain as compared with sustained weight problems. A significant energy lies in using a rigorous propensity rating matching technique, which minimized baseline confounding and enabled a balanced comparability between teams. In addition, the combination of scientific, biochemical, and liver tissue knowledge offered a singular alternative to look at hepatic inflammatory signatures related to weight regain, a context that continues to be insufficiently characterised in present literature. Beyond mechanistic concerns, early hepatic inflammatory alterations might have scientific implications in MASLD, a number one explanation for liver-related morbidity and mortality.⁴⁴ Macrophage activation, specifically, has been implicated as a central mediator linking metabolic stress to hepatic damage and illness development in MASLD.⁴⁵ In this context, the commentary of elevated transaminases and macrophage-related gene expression regardless of absence of fibrosis or superior histological modifications might symbolize a window of early hepatic vulnerability. While causality can’t be inferred, repeated episodes of weight regain may contribute to MASLD development by way of recurrent hepatic irritation.

Several limitations must be famous. First, mechanistic interpretations are derived from a male-predominant cohort. While this design diminished organic heterogeneity and strengthened inside validity, it limits the generalizability of our findings to feminine sufferers. Although all out there feminine contributors with liver biopsy samples are transparently reported (Supplementary Table 10), the small variety of feminine samples precluded statistically or biologically significant subgroup analyses; thus, whether or not comparable macrophage-associated inflammatory signatures exist in females stays an open query. Second, regardless of systematic EMR-based knowledge assortment, recall bias associated to long-term weight trajectories can’t be totally excluded, notably concerning the timing and magnitude of weight regain. Third, the cohort was restricted to people present process lifestyle-based weight reduction to keep away from heterogeneity launched by anti-obesity pharmacotherapies; consequently, the hepatic results of weight regain following pharmacological interventions stay unexplored. Finally, as a result of observational design, causal relationships can’t be established. Given the multifaceted and customized determinants influencing weight fluctuations,⁴⁶ it’s impractical to randomize contributors into the teams able to sustaining or regaining diminished physique weight. Therefore, the present observational examine might emphasize the need for designing research of upper degree of proof.

Conclusions

In conclusion, our examine gives novel proof that advantages achieved by means of lifestyle-induced weight reduction might not be sustained after subsequent weight regain. Compared with people who maintained a secure state of weight problems, these experiencing weight regain confirmed modest however constant elevations in hepatic transaminases and elevated hepatic expression of macrophage-related markers, regardless of the absence of superior steatosis or fibrosis. By integrating biochemical and liver tissue knowledge, our findings lengthen present data by indicating that weight regain might preferentially have an effect on hepatic inflammatory pathways earlier than overt metabolic dysfunction or structural liver abnormalities change into obvious.

Clinically, these outcomes recommend that reaching weight reduction alone could also be inadequate to make sure sturdy hepatic well being. Individuals with a historical past of weight regain following way of life interventions might profit from continued monitoring of liver-related biochemical parameters, even within the absence of established liver illness. Collectively, our findings underscore the significance of early liver threat evaluation in long-term weight administration and spotlight the necessity for longitudinal research to find out whether or not repeated weight regain contributes to progressive hepatic damage.

Abbreviations

MASLD, Metabolic dysfunction–related steatotic liver illness; PSM, Propensity rating matching; PCR, Polymerase chain response; ALT, Alanine aminotransferase; AST, Aspartate aminotransferase; DSI, Dallas Steatosis Index; NFS, Nonalcoholic fatty liver illness fibrosis rating; FIB-4, Fibrosis-4 index; NAS, NAFLD Activity Scores; NAFLD, Non-alcoholic related fatty liver illness; BMI, Body mass index; GOCY, Genetics of Obesity in Chinese Youngs; OGTT, Oral glucose tolerance take a look at; AKP, Alkaline phosphatase; GGT, γ-Glutamyl transpeptidase; TG, Triglyceride; TC, Total ldl cholesterol; HDL-C, High-density lipoprotein ldl cholesterol; LDL-C, Low-density lipoprotein ldl cholesterol; HbA1c, Glycosylated hemoglobin; fT3, Free triiodothyronine; fT4, Free tetraiodothyronine; TSH, Thyroid-stimulating hormone; HOMA-IR, Homeostasis mannequin evaluation of insulin resistance; HOMA-β, Homeostasis mannequin evaluation of β-cell operate; ISI, Insulin sensitivity index; DI, Disposition index; cDNA, Complementary deoxyribonucleic acid; PCA, Principal element evaluation; FDR, False discovery price; GSEA, Gene Set Enrichment Analysis; GO, Gene Ontology; IHC, Immunohistochemistry; BSA, Bovine serum albumin; IODs, Integrated optical densities; SMD, Standardized imply distinction; SD, Standard deviation; WBC, White blood cell; IQR, Interquartile vary; EMR, Electronic medical information; SEM, Standard error of the imply.

Data Sharing Statement

The datasets used and/or analyzed through the present examine can be found from the corresponding creator on cheap request.

Ethics Approval and Consent to Participate

Conduction of the examine together with the waiver of knowledgeable consent have been accepted by the Institutional Review Board of the Ruijin Hospital, Shanghai Jiao Tong University School of Medicine (Approval quantity: Ethics 2023[411]). Our examine complies with the Declaration of Helsinki.

Acknowledgments

The authors wish to thank Dr. Yuqin Cao from Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, for her useful assist and steerage within the statistical evaluation. The authors additionally lengthen their gratitude to all examine contributors and the scientific employees of Ruijin Hospital for his or her cooperation and contributions all through the analysis.

Author Contributions

Y.Z., Z.C. and Y.C. are co-first authors. Conceptualization: J.H., J.W., S.Z.; Methodology: Z.C., M.Y.; Investigation: Y.Z., Y.C., Z.Z., Z.C., M.Y.; Data Curation: Y.Z., Z.C., Y.C. W.G.; Formal Analysis: Y.Z., Y.C., Z.Z.; Resources: W.G., S.Z., J.H.; Supervision: J.H., J.W., S.Z., W.G.; Writing – Original Draft: J.H., Y.Z.; Writing – Review & Editing: all authors. All authors gave ultimate approval of the model to be revealed; have agreed on the journal to which the article has been submitted; and conform to be accountable for all elements of the work.

Funding

The design of the examine and assortment, evaluation, and interpretation of knowledge and writing the manuscript work have been supported by grants from National Key Research and Development Program of China (2023YFC2506001), the National Natural Science Foundation of China (82370845) and Natural Science Foundation of Shanghai (25ZR1402352).

Disclosure

The authors declare that they haven’t any competing pursuits.

References

1. Perdomo CM, Cohen RV, Sumithran P, Clément Okay, Frühbeck G. Contemporary medical, gadget, and surgical therapies for weight problems in adults. Lancet Lond Engl. 2023;401(10382):1116–15. doi:10.1016/S0140-6736(22)02403-5

2. European Association for the Study of the Liver (EASL). European Association for the Study of Diabetes (EASD), European Association for the Study of Obesity (EASO). EASL-EASD-EASO Clinical Practice Guidelines on the administration of metabolic dysfunction-associated steatotic liver illness (MASLD). J Hepatol. 2024;81(3):492–542. doi:10.1016/j.jhep.2024.04.031

3. Kanwal F, Neuschwander-Tetri BA, Loomba R, Rinella ME. Metabolic dysfunction-associated steatotic liver illness: replace and impression of recent nomenclature on the American Association for the Study of Liver Diseases apply steerage on nonalcoholic fatty liver illness. Hepatology. 2024;79(5):1212–1219. doi:10.1097/HEP.0000000000000670

4. Danpanichkul P, Suparan Okay, Diaz LA, et al. The Rising Global Burden of MASLD and Other Metabolic Diseases (2000-2021). United Eur Gastroenterol J. 2025;13(7):1141–1154. doi:10.1002/ueg2.70072

5. Targher G, Byrne CD, Tilg H. NAFLD and elevated threat of heart problems: scientific associations, pathophysiological mechanisms and pharmacological implications. Gut. 2020;69(9):1691–1705. doi:10.1136/gutjnl-2020-320622

6. Stefan N, Cusi Okay. A worldwide view of the interaction between non-alcoholic fatty liver illness and diabetes. Lancet Diabetes Endocrinol. 2022;10(4):284–296. doi:10.1016/S2213-8587(22)00003-1

7. Abdeldyem SM, Goda T, Khodeir SA, Abou Saif S, Abd-Elsalam S. Nonalcoholic fatty liver illness in sufferers with acute ischemic stroke is related to extra extreme stroke and worse end result. J Clin Lipidol. 2017;11(4):915–919. doi:10.1016/j.jacl.2017.04.115

8. Byrne CD, Targher G. NAFLD as a driver of persistent kidney illness. J Hepatol. 2020;72(4):785–801. doi:10.1016/j.jhep.2020.01.013

9. Malekpour MR, Abbasi-Kangevari M, Ghamari SH, et al. The burden of metabolic threat elements in North Africa and the Middle East, 1990-2019: findings from the Global Burden of Disease Study. EClinicalMedicine. 2023;60:102022. doi:10.1016/j.eclinm.2023.102022

10. Goodpaster BH, Delany JP, Otto AD, et al. Effects of weight loss program and bodily exercise interventions on weight reduction and cardiometabolic threat elements in severely overweight adults: a randomized trial. JAMA. 2010;304(16):1795–1802. doi:10.1001/jama.2010.1505

11. Dombrowski SU, Knittle Okay, Avenell A, Araújo-Soares V, Sniehotta FF. Long time period upkeep of weight reduction with non-surgical interventions in overweight adults: systematic evaluate and meta-analyses of randomised managed trials. BMJ. 2014;348(may14 6):g2646. doi:10.1136/bmj.g2646

12. Drucker DJ. Efficacy and Safety of GLP-1 Medicines for Type 2 Diabetes and Obesity. Diabetes Care. 2024;47(11):1873–1888. doi:10.2337/dci24-0003

13. Bessesen DH, Van Gaal LF. Progress and challenges in anti-obesity pharmacotherapy. Lancet Diabetes Endocrinol. 2018;6(3):237–248. doi:10.1016/S2213-8587(17)30236-X

14. Courcoulas AP, Christian NJ, Belle SH, et al. Weight change and well being outcomes at 3 years after bariatric surgical procedure amongst people with extreme weight problems. JAMA. 2013;310(22):2416–2425. doi:10.1001/jama.2013.280928

15. Courcoulas AP, King WC, Belle SH, et al. Seven-Year Weight Trajectories and Health Outcomes within the Longitudinal Assessment of Bariatric Surgery (LABS) Study. JAMA Surg. 2018;153(5):427. doi:10.1001/jamasurg.2017.5025

16. Yannakoulia M, Panagiotakos D. Weight loss by means of way of life modifications: impression within the major prevention of cardiovascular ailments. Heart. 2021;107(17):1429–1434. doi:10.1136/heartjnl-2019-316376

17. Kenneally S, Sier JH, Moore JB. Efficacy of dietary and bodily exercise intervention in non-alcoholic fatty liver illness: a scientific evaluate. BMJ Open Gastroenterol. 2017;4(1):e000139. doi:10.1136/bmjgast-2017-000139

18. Magkos F, Fraterrigo G, Yoshino J, et al. Effects of Moderate and Subsequent Progressive Weight Loss on Metabolic Function and Adipose Tissue Biology in Humans with Obesity. Cell Metab. 2016;23(4):591–601. doi:10.1016/j.cmet.2016.02.005

19. Wharton S, Lau DCW, Vallis M, et al. Obesity in adults: a scientific apply guideline. Can Med Assoc J. 2020;192(31):E875–91. doi:10.1503/cmaj.191707

20. Wing RR, Espeland MA, Clark JM, et al. Association of Weight Loss Maintenance and Weight Regain on 4-Year Changes in CVD Risk Factors: the Action for Health in Diabetes (Look AHEAD) Clinical Trial. Diabetes Care. 2016;39(8):1345–1355.

21. Liu M, Huang R, Xu L, et al. Cardiovascular results of intensive way of life intervention in adults with chubby/weight problems and sort 2 diabetes in accordance with physique weight time in vary. EClinicalMedicine. 2022;49:101451. doi:10.1016/j.eclinm.2022.101451

22. Bangalore S, Fayyad R, Laskey R, DeMicco DA, Messerli FH, Waters DD. Body-Weight Fluctuations and Outcomes in Coronary Disease. N Engl J Med. 2017;376(14):1332–1340. doi:10.1056/NEJMoa1606148

23. Zou H, Yin P, Liu L, et al. Body-Weight Fluctuation Was Associated With Increased Risk for Cardiovascular Disease, All-Cause and Cardiovascular Mortality: a Systematic Review and Meta-Analysis. Front Endocrinol. 2019;10:728. doi:10.3389/fendo.2019.00728

24. van Baak MA, Mariman ECM. Mechanisms of weight regain after weight reduction — the function of adipose tissue. Nat Rev Endocrinol. 2019;15(5):274–287. doi:10.1038/s41574-018-0148-4

25. Matsuda M, DeFronzo RA. Insulin sensitivity indices obtained from oral glucose tolerance testing: comparability with the euglycemic insulin clamp. Diabetes Care. 1999;22(9):1462–1470. doi:10.2337/diacare.22.9.1462

26. McHenry S, Park Y, Browning JD, Sayuk G, Davidson NO. Dallas Steatosis Index Identifies Patients With Nonalcoholic Fatty Liver Disease. Clin Gastroenterol Hepatol. 2020;18(9):2073–2080.

27. European Association for the Study of the Liver. Electronic handle: [email protected], Clinical Practice Guideline Panel, Chair:, EASL Governing Board consultant:, Panel members: EASL Clinical Practice Guidelines on non-invasive exams for analysis of liver illness severity and prognosis – 2021 replace. J Hepatol. 2021;75(3):659–689. doi:10.1016/j.jhep.2021.05.025

28. Sekhon JS. Multivariate and Propensity Score Matching Software with Automated Balance Optimization: the Matching Package for R. J Stat Softw. 2011;42(7). doi:10.18637/jss.v042.i07

29. Fan JG, Kim SU, Wong VWS. New tendencies on weight problems and NAFLD in Asia. J Hepatol. 2017;67(4):862–873. doi:10.1016/j.jhep.2017.06.003

30. Santos I, Sniehotta FF, Marques MM, Carraça EV, Teixeira PJ. Prevalence of non-public weight management makes an attempt in adults: a scientific evaluate and meta-analysis. Obes Rev off J Int Assoc Study Obes. 2017;18(1):32–50. doi:10.1111/obr.12466

31. MacLean PS, Wing RR, Davidson T, et al. NIH working group report: modern analysis to enhance upkeep of weight reduction. Obes Silver Spring Md. 2015;23(1):7–15. doi:10.1002/oby.20967

32. Jensen MD, Ryan DH, Apovian CM, et al. AHA/ACC/TOS guideline for the administration of chubby and weight problems in adults: a report of the American College of Cardiology/American Heart Association Task Force on Practice Guidelines and The Obesity Society. Circulation. 2014;129(25):S102–138. doi:10.1161/01.cir.0000437739.71477.ee

33. Thillainadesan S, Madsen S, James DE, Hocking SL. The impression of weight biking on well being outcomes in animal fashions: a scientific evaluate and meta‐evaluation. Obes Rev. 2022;23(5):e13416. doi:10.1111/obr.13416

34. List EO, Jensen E, Kowalski J, Buchman M, Berryman DE, Kopchick JJ. Diet-induced weight reduction is ample to scale back senescent cell quantity in white adipose tissue of weight-cycled mice. Nutr Healthy Aging. 2016;4(1):95–99. doi:10.3233/NHA-1614

35. Barbosa-da-Silva S, da Silva NC, Aguila MB, Mandarim-de-Lacerda CA. Liver injury will not be reversed through the lean interval in diet-induced weight biking in mice. Hepatol Res off J Jpn Soc Hepatol. 2014;44(4):450–459. doi:10.1111/hepr.12138

36. Kazankov Okay, Jørgensen SMD, Thomsen KL, et al. The function of macrophages in nonalcoholic fatty liver illness and nonalcoholic steatohepatitis. Nat Rev Gastroenterol Hepatol. 2019;16(3):145–159. doi:10.1038/s41575-018-0082-x

37. Thibaut R, Gage MC, Pineda‐Torra I, Chabrier G, Venteclef N, Alzaid F. Liver macrophages and irritation in physiology and physiopathology of non-alcoholic fatty liver illness. FEBS J. 2022;289(11):3024–3057. doi:10.1111/febs.15877

38. Ganz M, Bukong TN, Csak T, et al. Progression of non-alcoholic steatosis to steatohepatitis and fibrosis parallels cumulative accumulation of hazard alerts that promote irritation and liver tumors in a excessive fat-cholesterol-sugar weight loss program mannequin in mice. J Transl Med. 2015;13:193. doi:10.1186/s12967-015-0552-7

39. Carpino G, Nobili V, Renzi A, et al. Macrophage Activation in Pediatric Nonalcoholic Fatty Liver Disease (NAFLD) Correlates with Hepatic Progenitor Cell Response by way of Wnt3a Pathway. PLoS One. 2016;11(6):e0157246. doi:10.1371/journal.pone.0157246

40. Tada Y, Kasai Okay, Makiuchi N, et al. Roles of Macrophages in Advanced Liver Fibrosis, Identified Using a Newly Established Mouse Model of Diet-Induced Non-Alcoholic Steatohepatitis. Int J Mol Sci. 2022;23(21):13251. doi:10.3390/ijms232113251

41. Netea MG, Domínguez-Andrés J, Barreiro LB, et al. Defining educated immunity and its function in well being and illness. Nat Rev Immunol. 2020;20(6):375–388. doi:10.1038/s41577-020-0285-6

42. Cottam MA, Caslin HL, Winn NC, Hasty AH. Multiomics reveals persistence of obesity-associated immune cell phenotypes in adipose tissue throughout weight reduction and weight regain in mice. Nat Commun. 2022;13(1):2950. doi:10.1038/s41467-022-30646-4

43. Caslin HL, Cottam MA, Piñon JM, Boney LY, Hasty AH. Weight biking induces innate immune reminiscence in adipose tissue macrophages. Front Immunol. 2023;13:984859. doi:10.3389/fimmu.2022.984859

44. Powell EE, Wong VWS, Rinella M. Non-alcoholic fatty liver illness. Lancet. 2021;397(10290):2212–2224. doi:10.1016/S0140-6736(20)32511-3

45. Barreby E, Chen P, Aouadi M. Macrophage practical variety in NAFLD – greater than irritation. Nat Rev Endocrinol. 2022;18(8):461–472. doi:10.1038/s41574-022-00675-6

46. Varkevisser RDM, van Stralen MM, Kroeze W, Ket JCF, Steenhuis IHM. Determinants of weight reduction upkeep: a scientific evaluate. Obes Rev. 2019;20(2):171–211. doi:10.1111/obr.12772