This cross-sectional observational case-control study aimed to examined serum levels of leucine-rich repeat and fibronectin type III domain-containing protein 5 (LRFN5) and olfactomedin-4 (OLFM4) levels in methamphetamine use disorder (MUD), methamphetamine-induced psychotic disorder (MP), and healthy control (HC) groups. The study also aimed to assessed systemic inflammation using the Aggregate Index of Systemic Inflammation (AISI) and examined associations between biomarkers and clinical symptom severity, including Insight Assessment Scale (IAS) and Positive and Negative Syndrome Scale (PANSS) scores.

Methamphetamine is a potent central nervous system stimulant associated with addiction, neurotoxicity, and increased risk of psychosis. Chronic exposure affects dopaminergic systems in the mesolimbic and striatal pathways, potentially contributing to sensitization and psychotic symptom development. Methamphetamine-induced psychotic disorder (MP) occurs in a substantial proportion of users, yet individual vulnerability varies. Biological markers that may predict psychosis in methamphetamine users remain insufficiently studied.

Leucine-rich repeat and fibronectin type III domain-containing protein 5 (LRFN5) is a transmembrane synaptic adhesion molecule involved in synapse development, plasticity, and neuroimmune regulation. It contributes to excitatory and inhibitory synaptic differentiation and may influence neuroinflammatory responses. Olfactomedin-4 (OLFM4) is a neutrophil-specific glycoprotein involved in innate immunity, apoptosis regulation, inflammation, and glutamatergic synaptic modulation. These two markers are located in relatively close chromosomal regions and may demonstrate coordinated biological behavior.

To date, LRFN5 and OLFM4 have not been investigated in methamphetamine use disorder (MUD) or MP. This study is designed to compare serum levels of LRFN5 and OLFM4 among subjects with MP, subjects with MUD, and healthy controls (HC) subjects. A secondary objective was to evaluate associations between LRFN5, OLFM4 levels, psychotic symptom severity, and systemic inflammation, and to assess its potential predictive value for MP diagnosis.

Participants were planned to be consecutive inpatients diagnosed with MUD or MP according to the American Psychiatric Association's Fifth Edition Diagnostic and Statistical Manual of Mental Disorders (DSM-5-TR). Participant recruitment began on September 1, 2025, and is planned to be completed on April 1, 2026. All patients were selected/will be selected from those admitted to Elazığ Mental Health and Diseases Hospital. The HC group consisted of individuals without current or past psychiatric disorders or significant medical illnesses. Adult male participants aged 18-65 were included.

Venous blood samples were collected/will be collected at hospital admission prior to initiation of treatment. Samples were/will be centrifuged within 30 minutes and serum was stored at -80°C until analysis. Serum LRFN5 and OLFM4 levels were/will be measured using a commercially available enzyme-linked immunosorbent assay (ELISA) kit according to the manufacturer's instructions. Routine biochemical analyses and urine toxicology screening were/will be also performed. The Aggregate Index of Systemic Inflammation (AISI) was/will be calculated as (neutrophils × monocytes × platelets) / lymphocytes.

Psychotic symptom severity in the MP group was/will be assessed using the Positive and Negative Syndrome Scale (PANSS). Insight in the MP group was/will be assessed using the Insight Assessment Scale (IAS). Sociodemographic variables and clinical characteristics, including self-mutilation history, drug use onset age, duration of methamphetamine use, and smoking status were/will be recorded.

The study was/will be conducted in accordance with the Declaration of Helsinki and approved by the relevant institutional ethics committee. All participants provided/will provide written informed consent prior to participation.

1. For Methamphetamine Use Disorder (MUD) Group:

   *Inclusion Criteria:

   • Diagnosis of MUD according to DSM-5-TR
   • Medication-free for at least one month prior to admission
   • Age ≥ 18 years and <65 years
   • Provided informed consent

   For Methamphetamine Use Disorder (MUD) Group:

   *Exclusion Criteria:

   • Hypertension

   • Diabetes mellitus
   • Chronic kidney disease
   • Rheumatoid arthritis
   • Systemic lupus erythematosus
   • Cardiac illness
   • Severe neurological disorders
   • Immunological or systemic illness
   • Primary psychiatric disorders other than MUD
   • Alcohol use disorder

2. For Methamphetamine-Induced Psychotic Disorder (MP) Group:

   *Inclusion Criteria:

   • Diagnosis of MP according to DSM-5-TR
   • Medication-free for at least one month prior to admission
   • Age ≥ 18 years and <65 years
   • Provided informed consent

   For Methamphetamine-Induced Psychotic Disorder (MP) Group:

   *Exclusion Criteria:

   • Hypertension
   • Diabetes mellitus
   • Chronic kidney disease
   • Rheumatoid arthritis
   • Systemic lupus erythematosus
   • Cardiac illness
   • Severe neurological disorders
   • Immunological or systemic illness
   • Primary psychiatric disorders other than MP
   • Alcohol use disorder

3. For Healthy Control Group:

   *Inclusion Criteria:

   • No psychiatric diagnosis
   • No systemic or immunological illness
   • Medication-free for at least one month
   • Age ≥ 18 years and <65 years
   • Provided informed consent

   For Healthy Control Group:

   *Exclusion Criteria:

   • Hypertension

   • Diabetes mellitus
   • Chronic kidney disease
   • Rheumatoid arthritis
   • Systemic lupus erythematosus
   • Cardiac illness
   • Severe neurological disorders
   • Immunological or systemic illness
   • Primary psychiatric disorders other than schizophrenia
   • Alcohol/drug/substance use

The study population consisted/will be consisted of adult male inpatients aged 18-65 years who were admitted to Elazığ Mental Health and Diseases Hospital.

The MP group included/will include consecutive inpatients diagnosed with methamphetamine-induced psychotic disorder (MP) and the methamphetamine use disorder (MUD) group included/will include consecutive inpatients diagnosed with MUD according to the Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition, Text Revision (DSM-5-TR).

Both patient groups consisted of active methamphetamine users and were hospitalized for clinical management at the time of assessment. Urine toxicology screening was performed at admission to confirm substance use status.

The Healthy Control (HC) group consisted of adult male individuals without current or past psychiatric disorders and without substance use disorders, matched to patient groups in terms of age and body mass index. HCs had no significant medical illnesses.

Deidentified individual participant data (IPD) underlying the results reported in this study [including demographic variables, serum leucine-rich repeat and fibronectin type III domain-containing protein 5 (LRFN5) and Olfactomedin-4 (OLFM4) levels, complete blood count parameters, and Positive and Negative Syndrome Scale (PANSS) and Insight Assessment Scale (IAS)] will be made available to qualified researchers upon reasonable request for academic purposes.

Data will be shared after removal of all direct identifiers and in accordance with applicable ethical approvals and data protection regulations. Access to the data will require a methodologically sound research proposal and a data use agreement. Requests should be directed to the corresponding author.

Data will be available beginning 6 months after publication and will remain available for 5 years.

Access will be granted to researchers who provide a methodologically sound proposal. Requests must be approved by the principal investigator and may require a data use agreement in accordance with institutional and ethical regulations.