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Abstract5-aminolevulinic acid (ALA) is a non-protein amino acid that has been widely used in the fields of medicine and agriculture. This study aims to engineer the C5 pathway of the ALA biosynthesis in Escherichia coli BL21 to enhance ALA production. The ALA synthase genes gltX, hemA, and hemL were overexpressed in E. coli BL21 to lead to the increase in the production of ALA. The sRNA RyhB was also overexpressed to downregulate the expression of ALA dehydratase to reduce the downstream bioconversion of ALA to porphobilinogen. Next, the gene arcA was knocked out by CRISPR-Cas9 technology to open the TCA cycle to promote the respiratory metabolism of the strain to reduce the feedback inhibition of heme to ALA. The fermentation conditions of the engineered strain were optimized by response surface experiments. The time-course analysis of the ALA production was carried out in a 1 L shake flask. Through these efforts, the production of ALA in engineered strain reached 2953 mg/L in a 1 L shake flask. This study contributes to the industrial production of ALA by the engineered E. coli in the future. [ABSTRACT FROM AUTHOR]

Simple Summary: In brain tumor surgery, increasing the extent of resection has been shown to improve patient outcomes and survival. 5-ALA serves as a tumor visualization adjunct and has been approved for use in adult high-grade gliomas. The purpose of this systematic review was to assess whether 5-ALA has similar utility and safety in pediatric patients. A total of 249 pediatric cases were identified from the relevant literature, confirming the safety of 5-ALA in this population. Overall, the fluorescence rates and utility were favorable, although there were some variations across different tumor grades and types. While our preliminary findings suggest that 5-ALA is both safe and effective in pediatric brain tumor surgery, further systematic clinical studies are needed to validate these results. Background: 5-Aminolevulinic acid-guided surgery for adult gliomas has been approved by the European Medicines Agency and the US Food and Drug Administration, becoming a reliable tool for improving gross total resection rates and patient outcomes. This has led several medical centers to explore the off-label use of 5-ALA in the resection of pediatric brain tumors, assessing its efficacy and safety across various tumor types. However, given the differences between children and adults, the appropriateness of 5-ALA use in pediatric populations has not yet been fully established. Methods: We collected eligible publications from Embase, Scopus, PubMed, and Proquest, ultimately selecting 27 studies. Data extraction and retrospective analysis of 249 surgical cases were conducted to determine the current efficacy and safety of 5-ALA in pediatric brain tumors. The fluorescence rate and utility stratified by several clinical features, including WHO grade, tumor classification, and tumor location, were analyzed. Results: Most studies suggest that 5-ALA can enhance tumor identification in high-grade tumors, including glioblastomas and anaplastic astrocytomas. Changes in survival or recurrence rates associated with 5-ALA-guided resection have not been reported. None of the cases reported significant postoperative complications related to the use of 5-ALA. Conclusions: 5-ALA can aid in the resection of high-grade gliomas in pediatric patients. The efficacy of 5-ALA in low-grade gliomas and other tumors may require enhancement with additional tools or modified administration protocols. The safety of 5-ALA has reached a preliminary consensus, although further randomized controlled trials and data on survival and molecular characteristics are needed. [ABSTRACT FROM AUTHOR]

The widespread use of antibiotics in intensive animal husbandry, and the agricultural utilization of manure from such farms, imposes a significant burden on the environment. Consequently, the effects of antibiotics should be studied not only in animals and humans but also in all components of biocenoses and agrocenoses. In our study, we analyze the impact of four different concentrations of tetracycline present in soil (0, 5, 50, and 500 mg/kg of soil) on the growth and key photosynthesis parameters of pea seedlings: chlorophyll concentration, aminolevulinic acid concentration, aminolevulinic acid dehydrogenase activity, and ribulose bisphosphate carboxylase-oxygenase (RuBisCO) activity. At the lowest tetracycline concentration, chlorophyll content decreased by 13% compared to the control (0 tetracycline), while at the highest antibiotic concentration, it decreased by as much as 27%. Similarly, the decrease in aminolevulinic acid (a chlorophyll precursor) concentration was significant, amounting to 34%. However, the activity of the dehydrogenase enzyme, which consumes this precursor, decreased even more drastically by 51%, indicating significant disturbances in the light phase of photosynthesis. However, the activity of RuBisCO in pea plants subjected to tetracycline was even more severely affected, dropping by 58%, 69%, and 70% in soils with increasing concentrations of tetracycline. The reduction in enzyme activity could only partially be explained by a less pronounced decrease in the quantity of RuBisCO (large subunit) protein, which amounted to 6.5%, 11%, and 35% for tetracycline concentrations of 5, 50, and 500 mg/kg of soil, respectively. [ABSTRACT FROM AUTHOR]

Strawberry Fusarium wilt (SFW) is a systematic soil-borne disease caused by Fusarium oxysporum f. sp. fragaria (Fof), which infects the vascular bundles, blocking water and nutrient transport from roots to the aboveground. It is a severe pathogen which spreads rapidly and destroys strawberry production. Finding a way to control this disease is of great scientific value and practical importance. In this study, three fungi were isolated from the vascular tissues of sick strawberries in the field. After DNA sequencing, they were identified as Fof, Aspergillus fumigatus and Trichoderma harzianum, respectively, among which the first two are pathogens and the third is a probiotic. All fungi were controlled by thiophanate-methyl (TM), a commercial fungicide. On PDA medium, 20 mg•L-1 5-aminolevulinic acid (ALA), a natural non-protein amino acid, promoted T. harzianum proliferation, but inhibited Fof and A. fumigatus. In confrontation test, the growth of Fof or A. fumigatus was inhibited by T. harzianum and exogenous ALA promoted T. harzianum growth but significantly inhibited the pathogen growth. When three species of fungi were separately or combinedly inoculated on healthy strawberry plants, T. harzianum promoted plant growth and development while Fof or A. fumigatus caused growth retardation, where Fof directly caused leaf yellowing and plant wilting. When the plants inoculated with different fungus were treated with ALA, the results turned out that ALA alleviated SFW symptoms by bidirectionally promoting T. harzianum proliferation and inhibiting Fof and A. fumigatus. Thus, ALA might be used in comprehensively controlling SFW in strawberry industry. [ABSTRACT FROM AUTHOR]

Large, rapid extracellular oxygen transients (ΔpO 2) have been measured in vivo during ultra-high dose rate radiation therapy; however, it has been unclear if they match intracellular oxygen levels. Here, the endogenously produced protoporphyrin IX (PpIX) delayed fluorescence signal was measured as an intracellular in-vivo oxygen sensor to quantify these transients, with direct comparison to extracellular pO 2. Intracellular ΔpO 2 is closer to the cellular DNA, the site of major radiobiological damage, and therefore should help elucidate radiochemical mechanisms of the FLASH effect and potentially be translated to human tissue measurement. PpIX was induced in mouse skin through intraperitoneal injection of 250 mg/kg of aminolevulinic acid. The animals were also administered a 50 µL intradermal injection of 10 µM oxyphor G4 (PdG4) for phosphorescence lifetime pO 2 measurement. Paired oxygen transients were quantified in leg or flank tissues while delivering 10 MeV electrons in 3 µs pulses at 360 Hz for a total dose of 10 to 28 Gy. Transient reductions in pO 2 were quantifiable in both PpIX delayed fluorescence and oxyphor phosphorescence, corresponding to intracellular and extracellular pO 2 values, respectively. Reponses were quantified for 10, 22, and 28 Gy doses, with ΔpO 2 found to be proportional to the dose on average. The ΔpO 2 values were dependent on initial pO 2 in a logistic function. The average and standard deviations in ΔpO 2 per dose were 0.56 ± 0.18 mm Hg/Gy and 0.43 ± 0.06 mm Hg/Gy for PpIX and oxyphor, respectively, for initial pO 2 > 20 mm Hg. Although there was large variability in the individual animal measurements of ΔpO 2 , the average values demonstrated a direct and proportional correlation between intracellular and extracellular pO 2 changes, following a linear 1:1 relationship. A fundamentally new approach to measuring intracellular oxygen depletion in living tissue showed that ΔpO 2 transients seen during ultra-high dose rate radiation therapy matched those quantified using extracellular oxygen measurement. This approach could be translated to humans to quantify intracellular ΔpO 2. The measurement of these transients could potentially allow the estimation of intracellular reactive oxygen species production. [ABSTRACT FROM AUTHOR]

Different types of photosensitizers (PSs) have different dynamics and intensities of accumulation, depending on the type of tumor or different areas within the same tumor. This determines the effectiveness of fluorescence diagnostics and photodynamic therapy (PDT). This paper studies the processes of 5-aminolevulinic acid (5-ALA)-induced protoporphyrin IX (PpIX) and chlorin e6 (Ce6) accumulation in the central and border zones of a tumor after combined administration of two PSs into the patient's body. Fluorescence diagnostic methods have shown that sublingual administration of 5-ALA leads to the more intense accumulation of PpIX in a tumor compared to oral administration. Differences have been identified in the dynamics of 5-ALA-induced PpIX and Ce6 accumulation in the central and border zones of the tumor, as well as normal tissues. Ce6 accumulates mainly in the central zone of the tumor while PpIX accumulates in the border zone of the tumor. All patients with combined PDT experienced complete therapeutic pathomorphosis and relapse-free observation. [ABSTRACT FROM AUTHOR]

Background: Spine grape (Vitis davidii) is a promising source of high-quality anthocyanins, with vast potential for application in food, pharmaceutical, and cosmetic industries. However, their availability is limited by resource constraints. Plant cell culture has emerged as a valuable approach for anthocyanin production and serves as an ideal model to investigate the regulation of anthocyanin biosynthesis. Elicitors are employed to achieve targeted enhancement of anthocyanin biosynthesis. The present study investigated the impact of 5-aminolevulinic acid (ALA) as an elicitor on the accumulation of anthocyanins and flavonoids during spine grape callus growth. Specifically, we examined the effects of ALA on anthocyanin and its component accumulation in callus, and biosynthetic anthocyanin gene expression. Results: ALA at 25 µg/L increased the biomass of spine grape callus. ALA induction enhanced the levels of flavonoids, anthocyanins and proanthocyanidins in callus, with maximum values reaching 911.11 mg/100 g DW, 604.60 mg/100 g DW, and 5357.00 mg/100 g DW, respectively, after callus culture for 45 days. Notably, those levels were 1.47-, 1.93- and 1.83-fold higher than controls. ALA induction modulated the flavonoid profile, and among 97 differential flavonoid metabolites differing from controls, 77 were upregulated and 20 were downregulated. Six kinds of anthocyanins, namely cyanidin (8), delphinidin (6), peonidin (5), malvidin (4), petunidin (3) and pelargonidin (3), were detected in callus, with peonidin most abundant. Compared with controls, anthocyanin components were increased in ALA-treated callus. The key genes PAL1, PAL2, PAL4, CHI, CHS3, F3'H, F3H, FLS, DFR, UFGT, MYBA1, LDOX, OMT3, GT1 and ACT involved in anthocyanin biosynthesis were upregulated following ALA treatment, resulting in anthocyanin accumulation. Conclusion: This study revealed a novel mode of ALA-mediated promotion of plant anthocyanin biosynthesis and accumulation at the cellular level, and a strategy for enhancing anthocyanin content in spine grape callus. The findings advance commercial-scale production of anthocyanins via spine grape callus culture. we also explored the accumulation patterns of flavonoids and anthocyanins under ALA treatment. Augmentation of anthocyanins coincided with elevated expression levels of most genes involved in anthocyanin biosynthesis within spine grape callus following ALA treatment. [ABSTRACT FROM AUTHOR]

5-Aminolevulinic acid (5-ALA), a non-proteinogenic amino acid, is an intermediate in the biosynthesis of heme and exerts antiviral effects against feline coronavirus (FCoV); however, the underlying mechanisms remain unclear. In the biosynthesis of heme, 5-ALA is condensed and converted to protoporphyrin IX (PpIX), which is then transformed into heme by the insertion of ferrous iron. Previous research has suggested that the metabolites generated during heme biosynthesis contribute to the antiviral effects of 5-ALA. Therefore, the present study investigated the in vitro mechanisms responsible for the antiviral effects of 5-ALA. The results obtained revealed that 5-ALA and PpIX both effectively reduced the viral titer in the supernatant of FCoV-infected fcwf-4 cells. Moreover, PpIX exerted virucidal effects against FCoV. We also confirmed that 5-ALA increased PpIX levels in cells. While hemin induced heme oxygenase-1 gene expression, it did not reduce the viral titer in the supernatant. Sodium ferrous citrate decreased PpIX levels and suppressed the antiviral effects of 5-ALA. Collectively, these results suggest that the antiviral effects of 5-ALA against FCoV are dependent on PpIX. [ABSTRACT FROM AUTHOR]

This study evaluates the efficacy and safety of radiofrequency ablation (RFA) in treating low-grade squamous intraepithelial lesions (LSIL) with high-risk human papillomavirus (hrHPV) infection. The clinical data of 100 patients with LSIL and hrHPV infection were retrospectively analyzed. Patients were divided into the RFA group and the 5- aminolevulinic acid photodynamic therapy (ALA-PDT) group according to the treatment protocol, with 50 patients per group. Efficacy, negative hrHPV seroconversion, mental status, treatment satisfaction, and the occurrence of adverse reactions were compared between the two groups. The RFA group had a significantly higher total effective rate (94.00%) than the ALA-PDT group (74.00%). At 3 and 6-month follow-ups, the negative hrHPV seroconversion rate was significantly higher in the RFA group than in the ALA-PDT group (p < 0.05). At 6 months after treatment, the RFA group showed significantly lower Self-Rating Anxiety Scale scores and Self-Rated Depression Scale scores and higher satisfaction compared to the ALA-PDT group (all p < 0.05). The incidence of postoperative complications rate was not significantly different between the RFA group and ALA-PDT group (χ² = 3.184, p = 0.074). RFA is an effective and safe treatment that improves anxiety, depression, and satisfaction in patients with LSIL complicated by hrHPV infection. [ABSTRACT FROM AUTHOR]

Iron plays a crucial role in various metabolic processes. However, the impact of 5-aminolevulinic acid (ALA) in combination with iron chelators on iron metabolism and the efficacy of ALA-photodynamic therapy (PDT) remain inadequately understood. This study aimed to examine the effect of thiosemicarbazone derivatives during ALA treatment on specific genes related to iron metabolism, with a particular emphasis on mitochondrial iron metabolism genes. In our study, we observed differences depending on the cell line studied. For the HCT116 and MCF-7 cell lines, in most cases, the decrease in the expression of selected targets correlated with the increase in protoporphyrin IX (PPIX) concentration and the observed photodynamic effect, aligning with existing literature data. The Hs683 cell line showed a different gene expression pattern, previously not described in the literature. In this study, we collected an extensive analysis of the gene variation occurring after the application of novel thiosemicarbazone derivatives and presented versatile and effective compounds with great potential for use in ALA-PDT. [ABSTRACT FROM AUTHOR]

Background: Minimally-invasive 5-aminolevulinic acid photodynamic therapy (ALA-PDT) is used for treating cervical high-grade squamous intraepithelial lesions (HSIL/CIN2). The purpose of this study was to analyze the factors affecting the efficacy of ALA-PDT in the treatment of cervical HSIL/CIN2 in order to guide physicians in making appropriate treatment decisions. Methods: A retrospective study including 69 female patients with pathologically diagnosed HSIL/CIN2 was conducted. Patients were given six doses of 20% ALA-PDT at 7–14-day intervals. Cytology, HPV testing, colposcopy, and pathology were performed before treatment and at 6-month follow-up after treatment to assess efficacy. The main outcome of this study was the regression of HSIL/CIN2 and the clearance of high-risk HPV (hrHPV) infection after ALA-PDT treatment. Clinicopathological characteristics were collected to analyze the factors affecting the effectiveness of ALA-PDT treatment for HSIL/CIN2. Results: Between the successful and failed lesion regression group, there was a significant difference in sleeping disorders (p < 0.05). Between the successful and failed hrHPV clearance group, no statistically significant factors were found. With sensitivity values of 0.556 and 0.778, respectively, multivariate analysis showed that current smoking and sleeping disorders were independent prognostics of failure in lesion regression after ALA-PDT treatment. Conclusions: Smoking and sleep disorders were independent risk factors for failure in HSIL/CIN2 regression following ALA-PDT, suggesting the need for careful consideration of ALA-PDT for patients with these conditions. [ABSTRACT FROM AUTHOR]

Enhancing crop drought tolerance is crucial for food security amid climate change. This study examines how 5‐aminolevulinic acid (ALA) and hydrogen sulfide (H2S) can improve drought resilience in tomato plants, which are essential for sustainable food production. Drought stress was induced using 12% PEG‐6000. Plants were pre‐treated with 25 mg L−1 ALA and 0.1 mg L−1 hypotaurine (HT), followed by 0.2 mM sodium hydrosulfide (NaHS) treatment to assess the effects on plant physiological effects over 10 days. Drought stress reduced plant dry weight, chlorophylls (a and b), Fv/Fm, leaf water potential, and relative water content, while increasing glycine betaine (GB) and proline levels. Additionally, drought stress elevated NADPH oxidase (NOX) and glycolate oxidase (GOX) activities, inducing oxidative stress and membrane damage. ALA and NaHS enhanced plant growth, photosynthesis, proline, GB, ALA content, ATP synthase, and ATPase activities, while mitigating NOX and GOX activities, thereby reducing O2·−$$ {\mathrm{O}}_2^{\cdotp -} $$ and H2O2 radicals. ALA alone boosted L‐DES activity, promoting H2S accumulation. However, ALA + HT reduced H2S levels, compromising ALA's efficacy. NaHS with ALA + HT reinstated positive effects by restoring H2S levels. Biochemical assays confirmed ALA and NaHS promoted H2S accumulation, bolstering antioxidants, mitigating lipid peroxidation, suggesting their drought resilience potential in tomatoes. [ABSTRACT FROM AUTHOR]

BACKGROUND: In periprosthetic joint infections (PJIs), the surgeon's role becomes pivotal in addressing the infection locally, necessitating the surgical removal of infected and necrotic tissue. Opportunity to enhance the visualization of infected tissue during surgery could represent a game-changing innovation. OBJECTIVE: The aim of this narrative review is to delineate the application of intraoperative fluorescence imaging for targeting infected tissues in PJIs. METHODS: A systematic review, adhering to the Preferred Reporting Items for Systematic Review and Meta-Analyses (PRISMA) guidelines, was carried out. The search included multiple online database; MEDLINE, Scopus, and Web of Science. For data extraction the following were evaluated: (i) diagnosis of musculoskeletal infection; (ii) use of intraoperative fluorescence imaging; (iii) infected or necrotic tissues as target. RESULTS: Initially, 116 studies were identified through online database searches and reference investigations. The search was narrowed down to a final list of 5 papers for in-depth analysis at the full-text level. Subsequently, 2 studies were included in the review. The study included a total of 13 patients, focusing on cases of fracture-related infections of the lower limbs. CONCLUSION: The primary and crucial role for orthopedic surgeons in PJIs is the surgical debridement and precise removal of necrotic and infected tissue. Technologies that enable clear and accurate visualization of the tissue to be removed can enhance the eradication of infections, thereby promoting healing. A promising avenue for the future involves the potential application of intraoperative fluorescence imaging in pursuit of this objective. [ABSTRACT FROM AUTHOR]

Overexpression of recombinant Bacillus cereus TSPO (Bc TSPO) in E. coli bacteria leads to its recovery with a bound hemin both in bacterial membrane (MB) and inclusion bodies (IB). Unlike mouse TSPO, Bc TSPO purified in SDS detergent from IB is well structured and can bind various ligands such as high-affinity PK 11195, protoporphyrin IX (PPIX) and δ-aminolevulinic acid (ALA). For each of the three ligands, 1H–15N HSQC titration NMR experiments suggest that different amino acids of Bc TSPO binding cavity are involved in the interaction. PPIX, an intermediate of heme biosynthesis, binds to the cavity of Bc TSPO and its fluorescence can be significantly reduced in the presence of light and oxygen. The light irradiation leads to two products that have been isolated and characterized as photoporphyrins. They result from the addition of singlet oxygen to the two vinyl groups hence leading to the formation of hydroxyaldehydes. The involvement of water molecules, recently observed along with the binding of heme in Rhodobacter sphaeroides (Rs TSPO) is highly probable. Altogether, these results raise the question of the role of TSPO in heme biosynthesis regulation as a possible scavenger of reactive intermediates. [Display omitted] • Hemin is bound to Bc TSPO overexpressed in E. coli. • SDS-purified Bc TSPO is structured and binds ligands. • Purified Bc TSPO is functional and degrades PPIX upon UV light irradiation. • Oxygen-dependent PPIX degradation generates non-fluorescent products. • Products generated by ROS are hydroxyaldehydes. [ABSTRACT FROM AUTHOR]

Objective To explore the application of ALA photodynamic fluorescence combined with dermatoscopy in the diagnosis of facial cancerization and precancerous lesions. Methods From March 2022 to March 2024, a total of 52 patients with actinic keratosis, BOWEN's disease, squamous cell carcinoma and basal cell carcinoma were enrolled at the dermatology clinic of the First Affiliated Hospital of Henan University of Science and Technology. Each patient was examined by naked eye and ALA photodynamic fluorescence, and the difference in area was compared between these two methods. The ALA photodynamic fluorescence area was divided into 8 equal sections, followed by examination with a dermatoscope to determine whether there were cancerization and precancerous lesions. The lesion area and severity were scored. Results The ALA photodynamic fluorescence area was larger than that observed by the naked eye (all P

Abstract Background Spine grape (Vitis davidii) is a promising source of high-quality anthocyanins, with vast potential for application in food, pharmaceutical, and cosmetic industries. However, their availability is limited by resource constraints. Plant cell culture has emerged as a valuable approach for anthocyanin production and serves as an ideal model to investigate the regulation of anthocyanin biosynthesis. Elicitors are employed to achieve targeted enhancement of anthocyanin biosynthesis. The present study investigated the impact of 5-aminolevulinic acid (ALA) as an elicitor on the accumulation of anthocyanins and flavonoids during spine grape callus growth. Specifically, we examined the effects of ALA on anthocyanin and its component accumulation in callus, and biosynthetic anthocyanin gene expression. Results ALA at 25 µg/L increased the biomass of spine grape callus. ALA induction enhanced the levels of flavonoids, anthocyanins and proanthocyanidins in callus, with maximum values reaching 911.11 mg/100 g DW, 604.60 mg/100 g DW, and 5357.00 mg/100 g DW, respectively, after callus culture for 45 days. Notably, those levels were 1.47-, 1.93- and 1.83-fold higher than controls. ALA induction modulated the flavonoid profile, and among 97 differential flavonoid metabolites differing from controls, 77 were upregulated and 20 were downregulated. Six kinds of anthocyanins, namely cyanidin (8), delphinidin (6), peonidin (5), malvidin (4), petunidin (3) and pelargonidin (3), were detected in callus, with peonidin most abundant. Compared with controls, anthocyanin components were increased in ALA-treated callus. The key genes PAL1, PAL2, PAL4, CHI, CHS3, F3’H, F3H, FLS, DFR, UFGT, MYBA1, LDOX, OMT3, GT1 and ACT involved in anthocyanin biosynthesis were upregulated following ALA treatment, resulting in anthocyanin accumulation. Conclusion This study revealed a novel mode of ALA-mediated promotion of plant anthocyanin biosynthesis and accumulation at the cellular level, and a strategy for enhancing anthocyanin content in spine grape callus. The findings advance commercial-scale production of anthocyanins via spine grape callus culture. we also explored the accumulation patterns of flavonoids and anthocyanins under ALA treatment. Augmentation of anthocyanins coincided with elevated expression levels of most genes involved in anthocyanin biosynthesis within spine grape callus following ALA treatment.

Abstract Heme serves as a prosthetic group in hemoproteins, including subunits of the mammalian mitochondrial electron transfer chain. The first enzyme in vertebrate heme biosynthesis, 5-aminolevulinic acid synthase 1 (ALAS1), is ubiquitously expressed and essential for producing 5-aminolevulinic acid (ALA). We previously showed that Alas1 heterozygous mice at 20–35 weeks (aged-A1 +/−s) manifested impaired glucose metabolism, mitochondrial malformation in skeletal muscle, and reduced exercise tolerance, potentially linked to autophagy dysfunction. In this study, we investigated autophagy in A1 +/−s and a sarcopenic phenotype in A1 +/−s at 75–95 weeks (senile-A1 +/−s). Senile-A1 +/−s exhibited significantly reduced body and gastrocnemius muscle weight, and muscle strength, indicating an accelerated sarcopenic phenotype. Decreases in total LC3 and LC3-II protein and Map1lc3a mRNA levels were observed in aged-A1 +/−s under fasting conditions and in Alas1 knockdown myocyte-differentiated C2C12 cells (A1KD-C2C12s) cultured in high- or low-glucose medium. ALA treatment largely reversed these declines. Reduced AMP-activated protein kinase (AMPK) signaling was associated with decreased autophagy in aged-A1 +/−s and A1KD-C2C12s. AMPK modulation using AICAR (activator) and dorsomorphin (inhibitor) affected LC3 protein levels in an AMPK-dependent manner. Our findings suggest that heme deficiency contributes to accelerated sarcopenia-like defects and reduced autophagy in skeletal muscle, primarily due to decreased AMPK signaling.