Encircling granulosa cells protects against di-(2-ethylhexyl)phthalate-induced apoptosis in rat oocytes cultured in vitro.

The present study investigated if the presence of encircling granulosa cells protected against di(2-ethylhexyl)phthalate (DEHP)-induced oxidative stress in rat oocytes cultured in vitro. Denuded oocytes and cumulus-oocyte complexes (COCs) were treated with or without various doses of DEHP (0.0, 25.0, 50.0, 100, 200, 400 and 800 µM) in vitro. Morphological apoptotic changes, levels of oxidative stress and reactive oxygen species (ROS), mitochondrial membrane potential, and expression levels of apoptotic markers (Bcl2, Bax, cytochrome c) were analyzed. Our results showed that DEHP induced morphological apoptotic changes in a dose-dependent manner in denuded oocytes cultured in vitro. The effective dose of DEHP (400 µg) significantly (P>0.05) increased oxidative stress by elevating ROS levels and the mitochondrial membrane potential with higher mRNA expression and protein levels of apoptotic markers (Bax, cytochrome c). Encircling granulosa cells protected oocytes from DEHP-induced morphological changes, increased oxidative stress and ROS levels, as well as increased expression of apoptotic markers. Taken together our data suggested that encircling granulosa cells protected oocytes against DEHP-induced apoptosis and that the presence of granulosa cells could act positively towards the survival of oocytes under in vitro culture conditions and may be helpful during assisted reproductive technique programmes.

Synthetic phospholipids as specific substrates for plasma endothelial lipase.

We designed and prepared synthetic phospholipids that generate lyso-phosphatidylcholine products with a unique mass for convenient detection by LC-MS in complex biological matrices. We demonstrated that compound 4, formulated either as a Triton X-100 emulsion or incorporated in synthetic HDL particles can serve as a substrate for plasma EL with useful specificity.

Prolonged elevated postprandial sugar augments severity in kidney disease: a north Indian hospital-based study.

AIM: Diabetes plays a major role in progression of renal failure. The risk-factor profile changes during the progression of chronic kidney disease (CKD) from mild/moderate to end-stage renal disease. The relationship between glycemic indices, blood pressure, body mass index (BMI) and age at diagnosis in Indians has been less investigated. We assessed association of these risk factors with CKD stages in Indian population. METHODS: This study was carried out on patients (n = 162) who were diagnosed with CKD and normal control group (n = 155). For BMI, National Institutes for Health criteria were used to categorize the patients. RESULT: The mean age of CKD patients were significantly increased with the advancement of stage. BMI, systolic blood pressure (SBP), postprandial sugar level (PP), urea and creatinine were also significantly higher with elevated stages, whereas no differences were observed in diastolic blood pressure (DBP) and fasting blood sugar (FBS). The logistic regression study gave a significant result (p = 0.000) when we compared the group of CKD patients with established/prolonged postprandial blood sugar. It was independently associated with mild CKD [odds ratio (OR) = 5.213, 95% confidence interval (CI) = 2.06-13.21, p = 0.000], moderate CKD (OR = 7.724, 95% CI = 4.05-14.74, p = 0.000) and severe CKD (OR = 7.610, 95% CI = 4.03-14.36, p = 0.000). CONCLUSION: SBP and PP were the best predictors of prevalent nephropathy in this population, while DBP and FBS were found to be less effective. This may have implication for kidney disease risk stratification and protection.

Severe Acute Liver Injury: First Manifestation of Systemic Lupus Erythematosus.

Systemic lupus erythematosus (SLE) is a common multisystem disease characterised by a wide variety of presentation patterns and complex manifestations. As a lymphoid organ, the liver plays an important role in the immune response and is a target of autoimmune responses.1 SLE can affect the liver in approximately 25-60 % of patients during their disease course.2,3 Liver dysfunction and SLE can present with complicated differential diagnoses. Liver dysfunction in SLE is usually mild and rarely leads to advanced liver diseases such as cirrhosis and liver failure.4,5 Liver dysfunction in SLE is usually caused by non-SLE-related causes such as drug toxicity, fatty liver, alcoholism, and associated autoimmune hepatitis. However, primary liver involvement in SLE is also well-recognised. Patients with SLE who present with acute liver failure are rare. We report a rare case of SLE-associated acute severe liver injury along with a literature review.

Metabolic Reprogramming of Tumor-Associated Macrophages Using Glutamine Antagonist JHU083 Drives Tumor Immunity in Myeloid-Rich Prostate and Bladder Cancers.

Glutamine metabolism in tumor microenvironments critically regulates antitumor immunity. Using the glutamine-antagonist prodrug JHU083, we report potent tumor growth inhibition in urologic tumors by JHU083-reprogrammed tumor-associated macrophages (TAMs) and tumor-infiltrating monocytes. We show JHU083-mediated glutamine antagonism in tumor microenvironments induced by TNF, proinflammatory, and mTORC1 signaling in intratumoral TAM clusters. JHU083-reprogrammed TAMs also exhibited increased tumor cell phagocytosis and diminished proangiogenic capacities. In vivo inhibition of TAM glutamine consumption resulted in increased glycolysis, a broken tricarboxylic acid (TCA) cycle, and purine metabolism disruption. Although the antitumor effect of glutamine antagonism on tumor-infiltrating T cells was moderate, JHU083 promoted a stem cell-like phenotype in CD8+ T cells and decreased the abundance of regulatory T cells. Finally, JHU083 caused a global shutdown in glutamine-utilizing metabolic pathways in tumor cells, leading to reduced HIF-1α, c-MYC phosphorylation, and induction of tumor cell apoptosis, all key antitumor features. Altogether, our findings demonstrate that targeting glutamine with JHU083 led to suppressed tumor growth as well as reprogramming of immunosuppressive TAMs within prostate and bladder tumors that promoted antitumor immune responses. JHU083 can offer an effective therapeutic benefit for tumor types that are enriched in immunosuppressive TAMs.

A stimuli-responsive "smart probe" for selective monitoring of multiple-cations via differential analyses.

A dual-emissive benzimidazole-based ternary ruthenium complex containing multiple ionophores viz. carbonyl, amine, and imine exhibits discriminating behaviour for Pb(2+), Fe(2+) and Zn(2+), at parts-per-million concentrations, which leads to selective recognition of the above cations via differential responses using a single optical technique.

Epigenetic Silencing of p16INK4a gene in Sporadic Breast Cancer.

Epigenetic alterations of tumor suppressor genes (TSG) involved in the onset and progression of Breast Cancer (BC) may serve as biomarkers for early detection and prediction of disease prognosis. We have herein tried to determine the methylation status of TSG, p16INK4a, in our 50 BC patients and their association with clinicopathological parameters. The methylation status of the p16INK4a gene in fresh tissue samples from 50 patients with BC was assessed by methylation-specific polymerase chain reaction (MS-PCR). The mean age of BC patients was 49.30 ± 9.75 years. Of 50 BC samples tested, 21 (42%) had methylated p16INK4a gene. p16INK4a gene hypermethylation was significantly associated with age ≤ 50 years, premenopausal status and advanced BC stage. Multivariate analysis revealed a strong association between advanced BC stage (Stage III and Stage IV) and p16INK4a hypermethylation (P = 0.008, RR = 5.996, 95% CI = 1.581-22.739). p16INK4a methylation was significantly associated with Triple Negative BC (TNBC) (P = 0.045, OR = 4.181, 95% CI = 1.030-16.981). These findings indicate that p16INK4a hypermethylation frequently occurs in BC. Hypermethylation of p16INK4a in young, premenopausal, TNBC and with advance stage in BC patients suggests its association with aggressive BC.

MDA5 RNA-sensing pathway activation by Mycobacterium tuberculosis promotes innate immune subversion and pathogen survival.

Host cytosolic sensing of Mycobacterium tuberculosis (M. tuberculosis) RNA by the RIG-I-like receptor (RLR) family perturbs innate immune control within macrophages; however, a distinct role of MDA5, a member of the RLR family, in M. tuberculosis pathogenesis has yet to be fully elucidated. To further define the role of MDA5 in M. tuberculosis pathogenesis, we evaluated M. tuberculosis intracellular growth and innate immune responses in WT and Mda5-/- macrophages. Transfection of M. tuberculosis RNA strongly induced proinflammatory cytokine production in WT macrophages, which was abrogated in Mda5-/- macrophages. M. tuberculosis infection in macrophages induced MDA5 protein expression, accompanied by an increase in MDA5 activation as assessed by multimer formation. IFN-γ-primed Mda5-/- macrophages effectively contained intracellular M. tuberculosis proliferation to a markedly greater degree than WT macrophages. Further comparisons of WT versus Mda5-/- macrophages revealed that during M. tuberculosis infection MDA5 contributed to IL-1ß production and inflammasome activation and that loss of MDA5 led to a substantial increase in autophagy. In the mouse TB model, loss of MDA5 conferred host survival benefits with a concomitant reduction in M. tuberculosis bacillary burden. These data reveal that loss of MDA5 is host protective during M. tuberculosis infection in vitro and in vivo, suggesting that M. tuberculosis exploits MDA5 to subvert immune containment.

Virulence and pathogenicity determinants in whole genome sequence of Fusarium udum causing wilt of pigeon pea.

The present study deals with whole genome analysis of Fusarium udum, a wilt causing pathogen of pigeon pea. The de novo assembly identified a total of 16,179 protein-coding genes, of which 11,892 genes (73.50%) were annotated using BlastP and 8,928 genes (55.18%) from KOG annotation. In addition, 5,134 unique InterPro domains were detected in the annotated genes. Apart from this, we also analyzed genome sequence for key pathogenic genes involved in virulence, and identified 1,060 genes (6.55%) as virulence genes as per the PHI-BASE database. The secretome profiling of these virulence genes indicated the presence of 1,439 secretory proteins. Of those, an annotation of 506 predicted secretory proteins through CAZyme database indicated maximum abundance of Glycosyl hydrolase (GH, 45%) family proteins followed by auxiliary activity (AA) family proteins. Interestingly, the presence of effectors for cell wall degradation, pectin degradation, and host cell death was found. The genome comprised approximately 895,132 bp of repetitive elements, which includes 128 long terminal repeats (LTRs), and 4,921 simple sequence repeats (SSRs) of 80,875 bp length. The comparative mining of effector genes among different Fusarium species revealed five common and two specific effectors in F. udum that are related to host cell death. Furthermore, wet lab experiment validated the presence of effector genes like SIX (for Secreted in Xylem). We conclude that deciphering the whole genome of F. udum would be instrumental in understanding evolution, virulence determinants, host-pathogen interaction, possible control strategies, ecological behavior, and many other complexities of the pathogen.

Intravenous BCG vaccination reduces SARS-CoV-2 severity and promotes extensive reprogramming of lung immune cells.

Bacillus Calmette-Guérin (BCG) confers heterologous immune protection against viral infections and has been proposed as vaccine against SARS-CoV-2 (SCV2). Here, we tested intravenous BCG vaccination against COVID-19 using the golden Syrian hamster model. BCG vaccination conferred a modest reduction on lung SCV2 viral load, bronchopneumonia scores, and weight loss, accompanied by a reversal of SCV2-mediated T cell lymphopenia, and reduced lung granulocytes. BCG uniquely recruited immunoglobulin-producing plasma cells to the lung suggesting accelerated local antibody production. BCG vaccination also recruited elevated levels of Th1, Th17, Treg, CTLs, and Tmem cells, with a transcriptional shift away from exhaustion markers and toward antigen presentation and repair. Similarly, BCG enhanced recruitment of alveolar macrophages and reduced key interstitial macrophage subsets, that show reduced IFN-associated gene expression. Our observations indicate that BCG vaccination protects against SCV2 immunopathology by promoting early lung immunoglobulin production and immunotolerizing transcriptional patterns among key myeloid and lymphoid populations.

A case of "fresh rupture" after open repair of a ruptured Achilles tendon.

We present the case of Achilles tendon rupture in a 54-year-old man while rehabilitating after end-to-end open repair of an acute Achilles tendon rupture. Re-rupture after surgical repair of Achilles tendon is well known. The present case, however, is atypical, because the second rupture occurred significantly proximal to the first rupture. To our knowledge, this is the first time this has been described in English language studies. We have termed this incident a fresh rupture. A gastrocnemius turndown flap was used to repair the fresh rupture, which led to a satisfactory recovery. This case report serves to inform surgeons of the existence of this type of Achilles tendon rupture, while considering the possible etiologies and suggesting a technique that has been shown to be successful in the present case.

Nomogram of peripapillary retinal nerve fiber layer thickness in myopic eyes of north Indian population.

PURPOSE: To evaluate peripapillary-RNFL thickness in myopia by Cirrus OCT among north Indian population by spherical equivalent (SE), age, gender, and axial length (AL). METHODS: This was a cross-sectional study held during 2019-2020. Patients aged 18-60 years underwent ophthalmic examination including retinoscopy, AL, and OCT RNFL thickness. Persons with previous ocular surgery or ocular ailment other than refractive error were excluded. The peripapillary-RNFL thickness was noted and compared by demographic determinants. RESULTS: We examined 300 eyes of 300 persons (mean age: 30.75 ± 8.57 years; 144 males/156 females). Among them, 224 were myopes and 76 were emmetropes (EM). The mean SE was - 3.3 ± 0.4D (range: -11.0D to + 0.37D). The mean AL was 24.61 ± 1.92 mm (22.1-29.5). Overall temporal, nasal, superior, inferior, and mean peripapillary-RNFL thickness was 66.31 ± 7.58, 78.57 ± 16.00, 120.63 ± 11.69, 116.60 ± 15.80, and 95.50 ± 10.84 µm, respectively. Temporal, nasal, superior, inferior, and mean peripapillary-RNFL thickness was 73.97 ± 8.36, 94.84 ± 7.63, 127.96 ± 8.96, 136.89 ± 6.53, and 108.34 ± 6.28 µm, respectively, in EM eyes as compared to 63.71 ± 6.18, 73.05 ± 14.24, 118.21 ± 11.53, 109.71 ± 11.50, and 91.14 ± 8.31 µm, respectively, in myopic eyes (P < 0.001). Association of peripapillary-RNFL thickness with myopia and its different grades was P < 0.001. Association of mean peripapillary-RNFL thickness with age was P > 0.005 and gender was P = 0.168. Correlation between SE and RNFL thickness was positive and significant. Correlation between AL and RNFL thickness was negative but statistically significant. Association of AL with SE was P < 0.001. CONCLUSION: We provide normative peripapillary-RNFL thickness in the north Indian population in order to help in screening for myopia with comorbidity such as glaucoma based on RNFL thickness.

Reinforcement of nanoporous lanthanum-doped zinc borate by vanadium selenide nanosheets for improved tribological activity.

Nanoporous zinc borate (ZB) and 10% lanthanum-doped porous zinc borate (LZB) were synthesized to explore the role of porosity and doping in zinc borate during lubrication. HR-SEM, TEM, and HR-TEM authenticated nanoporous structures. The tribological properties of their blends with paraffin oil (PO) were compared by employing ASTM D4172 and ASTM D5183 norms on a four-ball tester. Vanadium selenide nanosheets (VSe2) were used to reinforce the structure of LZB for further advancement of the tribological properties. The superiority of the LZB/VSe2 over LZB and VSe2 nanosheets could be adjudged by tribological data. The porosity and lanthanum doping have yielded commendable tribological activity. The VSe2 nanosheets have strengthened the LZB matrix. The other constituent oxides of tribofilm from the LZB matrix, based on EDX analysis and XPS studies of the worn surface, ZnO, B2O3, La2O3, and V2O5, have abetted lubrication. The AFM and SEM investigations of wear track corroborated the tribological results.

Geochemical and Petrological Characterization of the Early Eocene Carbonaceous Shales: Implications for Oil and Gas Exploration in the Barmer Basin, Northwest India.

Carbonaceous shales of the Early Eocene Dharvi/Dunger Formation in the onshore Barmer Basin, northwest India were studied for the first time by integrating geochemical and organic petrological analyses. The carbonaceous shales of the Early Eocene Dharvi/Dunger Formation are characterized by a higher organic carbon content (TOC) of >10 wt % and consist mainly of a mixture of organic matter of types II and III kerogen, with exhibited hydrogen index values ranging between 202 and 292 mg HC/g TOC. The dominance of such kerogen is confirmed by the high amounts of huminite and fluorescent liptinite macerals. Consequently, the carbonaceous shales of the Early Eocene Dharvi/Dunger Formation are promising source rocks for both oil and gas generation potential, with oils of high wax contents, according to pyrolysis-gas chromatography results. The chemical and optical maturity results such as low values huminite/vitrinite reflectance, production index, and T max show that most of the examined carbonaceous shale rocks from the outcrop section of the Kapurdi mine have entered the low maturity stage of oil generation, exhibiting a range of immature to the very early-mature. Therefore, as highlighted in this study, the substantial abundance in hydrocarbon generation potential from these carbonaceous shales in the Dharvi/Dunger Formation may represent future conventional petroleum exploration in the southern part of the Barmer Basin, where the Dharvi/Dunger Formation has reached deeper burial depths.

Dynamic single-cell RNA sequencing reveals BCG vaccination curtails SARS-CoV-2 induced disease severity and lung inflammation.

COVID-19 continues to exact a toll on human health despite the availability of several vaccines. Bacillus Calmette Guérin (BCG) has been shown to confer heterologous immune protection against viral infections including COVID-19 and has been proposed as vaccine against SARS-CoV-2 (SCV2). Here we tested intravenous BCG vaccination against COVID-19 using the golden Syrian hamster model together with immune profiling and single cell RNA sequencing (scRNAseq). We observed that BCG reduced both lung SCV2 viral load and bronchopneumonia. This was accompanied by an increase in lung alveolar macrophages, a reversal of SCV2-mediated T cell lymphopenia, and reduced lung granulocytes. Single cell transcriptome profiling showed that BCG uniquely recruits immunoglobulin-producing plasma cells to the lung suggesting accelerated antibody production. BCG vaccination also recruited elevated levels of Th1, Th17, Treg, CTLs, and Tmem cells, and differentially expressed gene (DEG) analysis showed a transcriptional shift away from exhaustion markers and towards antigen presentation and repair. Similarly, BCG enhanced lung recruitment of alveolar macrophages and reduced key interstitial macrophage subsets, with both cell-types also showing reduced IFN-associated gene expression. Our observations indicate that BCG vaccination protects against SCV2 immunopathology by promoting early lung immunoglobulin production and immunotolerizing transcriptional patterns among key myeloid and lymphoid populations.

124I-Iodo-DPA-713 Positron Emission Tomography in a Hamster Model of SARS-CoV-2 Infection.

PURPOSE: Molecular imaging has provided unparalleled opportunities to monitor disease processes, although tools for evaluating infection remain limited. Coronavirus disease (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is mediated by lung injury that we sought to model. Activated macrophages/phagocytes have an important role in lung injury, which is responsible for subsequent respiratory failure and death. We performed pulmonary PET/CT with 124I-iodo-DPA-713, a low-molecular-weight pyrazolopyrimidine ligand selectively trapped by activated macrophages cells, to evaluate the local immune response in a hamster model of SARS-CoV-2 infection. PROCEDURES: Pulmonary 124I-iodo-DPA-713 PET/CT was performed in SARS-CoV-2-infected golden Syrian hamsters. CT images were quantified using a custom-built lung segmentation tool. Studies with DPA-713-IRDye680LT and a fluorescent analog of DPA-713 as well as histopathology and flow cytometry were performed on post-mortem tissues. RESULTS: Infected hamsters were imaged at the peak of inflammatory lung disease (7 days post-infection). Quantitative CT analysis was successful for all scans and demonstrated worse pulmonary disease in male versus female animals (P < 0.01). Increased 124I-iodo-DPA-713 PET activity co-localized with the pneumonic lesions. Additionally, higher pulmonary 124I-iodo-DPA-713 PET activity was noted in male versus female hamsters (P = 0.02). DPA-713-IRDye680LT also localized to the pneumonic lesions. Flow cytometry demonstrated a higher percentage of myeloid and CD11b + cells (macrophages, phagocytes) in male versus female lung tissues (P = 0.02). CONCLUSION: 124I-Iodo-DPA-713 accumulates within pneumonic lesions in a hamster model of SARS-CoV-2 infection. As a novel molecular imaging tool, 124I-Iodo-DPA-713 PET could serve as a noninvasive, clinically translatable approach to monitor SARS-CoV-2-associated pulmonary inflammation and expedite the development of novel therapeutics for COVID-19.

Obesity augmented breast cancer risk: a potential risk factor for Indian women.

BACKGROUND: Obesity is caused by disturbances of energy balance, which is homeostasized by the physiological processes. The study aims to determine the possible impact of rising prevalence of obesity and its effect in the development of breast carcinoma (BC) in Indian population. METHODS: This study is carried out on patients (N = 358) who were diagnosed with BC and breast diseases (BD) by calculating their BMI admitted during the period of 2005 to 2009. NIH criteria were used to categorize the patients. Pathological factors of BC patient were then compared among groups. RESULT: These results were indicative of significant positive association between BC risks with peri/post menopausal status, residence, diet nature, and tobacco uses. Metastases were identified more commonly with increasing weight. It was found to be independently associated with obesity I (OR = 3.103, 95% CI = 1.633-5.895) and obesity II (OR = 6.803, 95% CI = 2.415-19.162). Disease stage and cancer related mortality were significantly associated with increased BMI. CONCLUSION: The higher prevalence of severe obesity among Indian population was associated with BC. The only alteration apart from early diagnosis is opting for a more natural lifestyle that will affect energy equilibrium and prove to be a viable option for prevention in carcinoma of breast for better survival.

BCG turns 100: its nontraditional uses against viruses, cancer, and immunologic diseases.

First administered to a human subject as a tuberculosis (TB) vaccine on July 18, 1921, Bacillus Calmette-Guérin (BCG) has a long history of use for the prevention of TB and later the immunotherapy of bladder cancer. For TB prevention, BCG is given to infants born globally across over 180 countries and has been in use since the late 1920s. With about 352 million BCG doses procured annually and tens of billions of doses having been administered over the past century, it is estimated to be the most widely used vaccine in human history. While its roles for TB prevention and bladder cancer immunotherapy are widely appreciated, over the past century, BCG has been also studied for nontraditional purposes, which include (a) prevention of viral infections and nontuberculous mycobacterial infections, (b) cancer immunotherapy aside from bladder cancer, and (c) immunologic diseases, including multiple sclerosis, type 1 diabetes, and atopic diseases. The basis for these heterologous effects lies in the ability of BCG to alter immunologic set points via heterologous T cell immunity, as well as epigenetic and metabolomic changes in innate immune cells, a process called "trained immunity." In this Review, we provide an overview of what is known regarding the trained immunity mechanism of heterologous protection, and we describe the current knowledge base for these nontraditional uses of BCG.

Recombinant BCGs for tuberculosis and bladder cancer.

Bacillus Calmette-Guérin (BCG) vaccine is an attenuated live strain of Mycobacterium bovis. It may be the most widely used vaccine in human history and is the only licensed human tuberculosis (TB) vaccine available. Despite its excellent safety history, a century of use in global vaccination programs, and its significant contribution to reducing TB mortality among children, the efficacy of BCG continues to be disputed due to its incomplete protection against pulmonary TB in adults. Still vaccines offer the best chance to contain the ongoing spread of multi-drug resistance TB and disease dissemination. The development of improved vaccines against TB therefore remains a high global priority. Interestingly, recent studies indicate that genetically modified BCG, or administration of existing BCG through alternate routes, or revaccination, offers improved protection, suggesting that BCG is well poised to make a comeback. Intravesical BCG is also the only approved microbial immunotherapy for any form of cancer, and is the first-line therapy for treatment-naïve non-muscle invasive bladder cancer (NMBIC), which represents a majority of the new bladder cancer cases diagnosed. However, almost a third of patients with NMIBC are either BCG unresponsive or have tumor recurrence, leading to a higher risk of disease progression. With very few advances in intravesical therapy over the past two decades for early-stage disease, and a limited pipeline of therapeutics in Phase 3 or late Phase 2 development, there is a major unmet need for improved intravesical therapies for NMIBC. Indeed, genetically modified candidate BCG vaccines engineered to express molecules that confer stronger protection against pulmonary TB or induce potent anti-tumor immunity in NMIBC have shown promise in both pre-clinical and clinical settings. This review discusses the development of second generation, genetically modified BCG candidates as TB vaccines and as anti-tumor adjuvant therapy for NMIBC.

Tetrahydropyrazolopyridines as antifriction and antiwear agents: experimental and DFT calculations.

Some tetrahydropyrazolopyridines (THPP-H) with the methoxy (THPP-OMe) and methyl (THPP-Me) substituents were synthesized by a one-pot multi-component reaction. NMR spectroscopy (1H and 13C) was used to authenticate the synthesis. According to the results of tribological tests ASTM D4172, and ASTM D5183 on a four-ball tester in paraffin oil (PO) at a concentration of 0.25% w/v, their relative tribo-activity along with a reference additive, zinc dialkyldithiophosphate (ZDDP) could be figured out as mentioned below-THPP-OMe > THPP-Me > THPP-H > ZDDP. The calculation of frictional power loss from the coefficient of friction data of the tested additives supports the given order. As is apparent from AFM and SEM micrographs of the wear scar surface for plain oil with and without different tetrahydropyrazopyridines, surface evenness endorses the above trend. Proof for strong adsorption of the synthesized additives is provided by EDX analysis of the steel ball surface after performing the tribological test, where nitrogen and oxygen are vividly seen as heteroatoms. XPS studies reveal the composition of the in situ formed tribofilm. The moieties containing carbon bonded to oxygen/nitrogen as decomposed products of the additive together with oxides of iron in +II or +III oxidation states are perceptible in the tribofilm, the tribofilm interferes with the proximity of the surfaces keeping them far apart. Consequently, friction and wear are remarkably reduced. Findings from Density Functional Theory (DFT) calculations are in full agreement with the results obtained from tribological experiments.