In vitro prebiotic effects of seaweed polysaccharides

Although prebiotic activities of alginate and agar oligosaccharides isolated from seaweeds have been reported, it remains unknown whether seaweed polysaccharides have prebiotic activity. In this study, we isolated polysaccharides from four species of seaweeds, such as Grateloupia fi licina(GFP), Eucheuma spinosum(ESP), Ulva pertusa(UPP), and A scophyllum nodosum(ANP), and characterized their structures and prebiotic ef fects in vitro. The results showed that these polysaccharides were dif ferent in total sugar and sulfate contents as well as monosaccharide composition. GFP and ESP significantly promoted bifi dobacterium proliferation and 0.1% ESP and 0.4% GFP resulted in the highest proliferation rates of beneficial bacteria, whereas UPP and ANP inhibited the growth of beneficial bacteria at all tested concentrations(0.1%–0.5%). The different behaviors of the four seaweed-originated polysaccharides might be refl ected by differences in monosaccharide composition and structure. Therefore, polysaccharides isolated from GFP and ESP could be utilized as prebiotics. However, more studies must be carried out in vivo.

The Inhibitory Effect of Extracts From Fructus lycii and Rhizoma polygonati on in vitro DNA Breakage by Alternariol

Alternariol caused DNA single-strand breakage. Conversion of the closed circular double-stranded supercoiled DNA (pBR 322) to the nicked circular form and linear form was used to investigate the effect of extracts of some Chinese medical herbs on DNA nicking induced by alternariol. Some substances in the extracts of Rhizoma polygonati (RP) and Fructus lycii (FL) were shown to protect DNA from the attack by alternariol.Some substance in the RP may bind to plasmid DNA, and this binding reduces the electrophoretic mobility of DNA. These results indicate that substances from FL and RP may be used as DNA protectors. It is possible that they play an important role in preventing cancer.

The Blocking Effect of Phenolic Acid on N-Nitrosomorpholine Formation in vitro

Phenolic acids (PAs) are widely found in many daily consumed vegetables and fruits.The inhibitory effects of PAs on Nnitrosomorpholine (NMOR) formation in vitro undersimulated gastric juice condition were studied. The results showed that the inhibjtoryIX)tency of thirteen varieties of PAs differed greatly, which may be related to theirchemical structures; the blocking rate (BR) of different killds of PAs were as follows:caf feic acid, 92.5%; tannic acid, 90.0%; gallic acid, 86.8%; sinaPinic acid, 86.2%; ferulicacid, 8l.l%; chlorogenic acid, 69.4%; gentisic acid, 69.2%; syringic acid, 62.1%; protocatechuic acid, 56.0%; p-coumaric acid, 52.5%; vannilic acid, 35.4%; moreover,p-hydroxybenzoic acid and m-coumaric acid had the least blocking effect or even slightcatalyzing effect. The results also demonstrated that amounts of NMOR formed werenegatively correlated with molar ratio of PAs to nitrite and that the optimum pH forinhibition was between 2 and 3.

THE SPECIFIC PHOTODYNAMIC EFFECTS OF MONOCLONAL ANTIBODIES CONJUGATED WITH HEMATOPORPHYRIN DERIVATIVE ON GASTRIC CANCER IN VITRO AND IN VIVO

Murine monoclonal antibody (MoAb) BB4.3, raised against the human gastric cancer cell line BGC823, was puriffied with Protein A-Sepharose CL-4B affinity chromatography and identified as IgG2a. It was then conjugated with a hematoporphyrin derivative (HPD) by using carbodiimide. The qualitative analysis of this conjugate showed that the amount of free HPD was negligible and there were no IgG aggregates among the conjugates. The conjugate retained both the antibody and photochemical activity of HPD.In vitro, the phototoxic effect of this HPD-BB4.3 conjugate on target cells was about 15 times higher than that of free HPD. The quality of selective photocytotoxicity was proven by the greater cytotoxi-city this conjugate showed than that of corresponding normal mouse IgG (NIgG) conjugated with HPD. It showed less cytotoxicity to colon cancer cell line B-80 (negative reaction to MoAb BB4.3) than to BGC825. Moreover, its cytotoxicity to BGC823 cells could be blocked specifically by excess BB4.3 antibody, but not by another MoAb 3G9, which combines with BGC823 at different binding sites from MoAb BB4.3.Nude mice inoculated with 2 × 10- BGC823 cells were given HPD-BB4.3, HPD, HPD-NIgG, HPD plus BB4.3 and PBS, respectively then exposed to light. Four out of six animals treated with the HPD-BB4.3 conjugate remained tumor-free for a long period. Although two developed tumors, there was a significant difference between the HPD-BB4.3-treated group and all the control groups in tumor induction time, tumor growth rate, and survival time (p<0.001). The HPD-BB4.3 conjugate inhibited the growth of established tumors by more than 40% in comparison with control groups (p<0.05).

ANTI-HUMAN LUNG GIANT CELL CANCER (PG) EFFECT OF HUMAN LAK CELLS IN VITRO AND IN NUDE MICE

Human LAK cells were prepared by culturing normal human peripheral blood mononuclear cells (PBMC) with or without rIL-2 and assayed for T cell surface markers as well as anti-tumor activity against PC in vitro and in nude mice. Although the percentages of T3, T4, and T8 positive cells in rIL-2-activated cells did not differ significantly from those of control cells in vitro, the former showed stronger cytotoxicity than control cells to PG tumor cells in vitro. In vivo, LAK cells completely inhibited the growth of PG tumor in nude mice, whereas PBMC control cells were to be of no effect. The anti-tumor effect of human LAK cells in nude mice may offer a useful model to study the role of human LAK cells against human tumor in vivo.

IN VITRO ANTI-HEPATOMA EFFECTS OF MONOCYTES AND KUPFFER CELLS ISOLATED FROM HEPATOMA PATIENTS AFTER TREATMENT WITH BIOLOGICAL IMMUNE STIMULANTS

Monocytes (MC), lymphocytes (LC) and Kupffer cells (KC) were isolated respectively from blood and surgical liver samples of patients suffering from he-patocellular carcinoma (HCC). 13 patients were given BCG, mixed bacterium vaccine (MBV) and human white blood cell interferon (IFN), the other 3 patients were not treated with any biological immune stimulants (BIS) and served as controls. The cytosta-tic and cytotoxic effects of MC and KC on human hepatoma SMMC-7721 (TC) were assayed in vitro and the numbers of T total (Tt), T helper (Th) and T suppressor (Ts) cells were counted using CD monoclonal antibody immunofluorescence. The results were as follows: (1) On the 7th day after the first administration of BIS, the cytostatic and cytotoxic effects of MC on TC showed obvious increase over pre-administration. The activity of BIS was 1 ?5 times as high as that in the controls. (2) After 3 administrations, the cytostatic effect of MC on TC increased to the normal level (84%), while the controls remained as before (45%). (3) On the 7th day after first administration, cytostatic and cytotoxic effects of KC on TC were 0.5 and 1 times higher respectively than those of the controls. (4) The numbers of Tt and Th of patients given BIS increased continuously; on the contrary Ts decreased in number. These results indicate that combined use of BCG, MBV and IFN can actively enhance the immune anti-hepatoma function of patients suffering from HCC.

The Effects of Dietary Restriction and Aging on in Vivo and in Vitro Binding of Aflatoxin B_1 to Cellular DNA

Laboratory animals maintained on a reduced calorie but nutritionally adequate diet have extended life spans and lowered incidences of spontaneous and chemically induced cancers compared to ad libitum- fed counterparts. Many of the effects of dietary restriction on laboratory animals have been suggested to be related to a deceleration of the aging process. The inhibition of age-related changes in xenobiotic metabolizing enzyme activities by dietary restriction has previously been reported. Alterations of these enzyme activities may cause changes in metabolic activation of carcinogens and, therefore, carcinogen-DNA binding. DNA-repair capability has also been reported to be enhanced in diet-restricted rats. Using AFB1 as a model carcinogen, we have studied in vivo and in vitro hepatic AFB1 -DNA binding, demonstrating that dietary restriction (60% of ad libitum consumption) may decrease the metabolic activation of AFB1, and subsequently reduce AFB 1-DNA binding. Our preliminary results obtained from the AFB 1-DNA binding experiments in isolated hepatocytes suggest that the observed age-dependent reduction in AFB 1-DNA binding which may be attributed to a loss of metabolic activating capability was delayed in the diet-restricted rats.

Effect of polygonatum polysaccharide on the hypoxia-induced apoptosis and necrosis in in vitro cultured cerebral cortical neurons from neonatal rats

BACKGROUND： Cardiocerebrovascular diseases induced cerebral circulation insufficiency and senile vascular dementia can result in ischemic/hypoxic apoptosis of central neurons, which we should pay more attention to and prevent and treat as early as possible. Traditional Chinese medicine possesses the unique advantage in this field. Polygonatum, a Chinese herb for invigorating qi, may play a role against the hypoxic apoptosis of brain neurons. OBJECTIVE ： To observe the protective effect of polygonatum polysaccharide on hypoxia-induced apoptosis and necrosis in cerebral cortical neurons cultured in vitro. DESIGN： A comparative experiment.SETTING： Laboratory of Cell Biology, Institute of Basic Medical Sciences, Jiangxi Provincial Academy of Traditional Chinese Medicine. MATERIALS： The experiment was carried out in the Laboratory of Cell Biology, Institute of Basic Medical Sciences, Jiangxi Provincial Academy of Traditional Chinese Medicine from November 2003 to April 2005. Totally 218 Wistar rats （male or female） of clean degree within 24 hours after birth were purchased from the animal center of Jiangxi Medical College （certification number was 021-97-03）. METHODS：① Preparation of cerebral cortical neurons of rats： The cerebral cortical tissues were isolated from the Wistar rats within 24 hours after birth, and prepared to single cell suspension, and the cerebral cortical neurons of neonatal rats were in vitro cultured in serum free medium with Neurobasal plus B27 Supplement. ② Observation on the non-toxic dosage of polygonatum polysaccharide on neurons： After the neurons were cultured for 4 days, polygonatum polysaccharide of different dosages （1-20 g/L） was added for continuous culture for 48 hours, the toxicity and non-toxic dosage of polygonatum polysaccharide on neurons were observed and detected with trypan blue staining. ③Grouping： After hypoxia/reoxygenation, the cultured neurons were divided into normal control group, positive apoptotic group and polygonatum polysaccharide group. In the normal control group, the neurons were cultured at 37℃ in CO2 with the volume fraction of 0.05 under saturated humidity for 6 days. In the apoptotic positive group, the neurons were cultured with hypoxia for 12 hours after 4-day culture, and followed by reoxygenation for 48 hours. In the polygonatum polysaccharide group, polygonatum polysaccharide with the terminal concentration of 0.5, 1 and 1.5 g/L was added to some neurons at 10 hours before the hypoxia culture, and then the neurons were cultured with hypoxia for 12 hours, followed by reoxygenation for 48 hours; polygonatum polysaccharide with the terminal concentration of 0.5, 1 and 1.5 g/L was added to the other neurons at 12 hours after hypoxia followed by reoxygenation for 48 hours.④ The Hoechst33342 fluorescence staining, Annexin V/PI flow cytometer, appearance of DNA agarose gel electrophoresis gradient strap and immunohistochemical staining were used to observe the expressions of Bcl-2, Bax and Caspase-3 apoptotic and anti-apoptotic proteins and the ratio of Bcl-2/Bax, and observe the effect of polygonatum polysaccharide against the hypoxic apoptosis of cerebral cortical neurons of neonatal rats. MAIN OUTCOME MEASURES： ① Toxicity and non-toxic dosage of polygonatum polysaccharide on neurons;② Apoptotic rate of neurons detected with Hoechst33342 fluorescence staining;③ Early apoptotic rate and necrotic rate of neurons detected with Annexin V/PI flow cytometer; ④DNA agarose gel electrophoresis ladder-like strap appeared or not;⑤ Expressions of Bcl-2, Bax and Caspase-3 apoptotic and anti-apoptotic proteins and the ratio of Bcl-2/Bax. RESULTS：① Polygonatum polysaccharide within 6 g/L had no cytotoxicity on the normal cultured cerebral cortical neurons （P 〉 0.05）. ②The apoptotic rates of neurons detected with Hoechst33342 fluorescence staining had significant differences between the polygonatum polysaccharide groups and positive apoptosis group added to neurons at 10 hours before the hypoxia culture [（13.00±4.52）%,（12.72±2.15）%, （11.80±1.18）%,（38.03±1.05）%, P 〈 0.01], and had no significant differences between the polygonatum polysaccharide groups and positive apoptosis group added to neurons at 12 hours after the hypoxia culture （36.77±1.45）%, （36.60±1.61）%, （36.37±2.02）%, （38.03±1.05）%, P 〉 0.05].③ Annexin V/PI flow cytometer detected that the anti-necrotic effect was enhanced with the increased concentration of polygonatum polysaccharide within 0.5-1.5 g/L （P 〈 0.01）. Polygonatum polysaccharide of 0.5-1.5 g/L added before hypoxia could significantly decrease the apoptotic rate of neurons induced by hypoxia/reoxygenation （P 〈 0.01）. ④ No DNA agarose gel electrophoresis ladder-like strap appeared in the groups with polygonatum polysaccharide of 0.5-1.5 g/L added at 10 hours before hypoxia;⑤ After Polygonatum polysaccharide of 0.5-1.5 g/L was added before hypoxia, the expression of Bcl-2 protein of hypoxic neurons was increased （P 〈 0.01）, and those of Bax protein and Caspase-3 protein were reduced （P 〈 0.01）, and the ratio of Bcl-2/Bax was increased （P 〈 0.01）. CONCLUSION： Polygonatum polysaccharide within 6 g/L has no cytotoxicity on the normal cultured cerebral cortical neurons. Polygonatum polysaccharide of 0.5-1.5 g/L added before hypoxia plays a role agains necrosis of neurons induced by hypoxia. Polygonatum polysaccharide of 0.5-1.5 g/L can significantly reduce the apoptosis of neurons induced by hypoxia through up-regulating the expression of Bcl-2 protein, down-regulating the expressions of Bax protein and Caspase-3 protein, and increasing the ratio of Bcl-2/Bax.

Effect of temsirolimus on bladder cancer cells in vitro and in vivo

Objective To examine the effects of temsirolimus, an inhibitor of mammalian target of rapamycin,on bladder cancer cell lines T24 and BIU-87 in vitro and in vivo for purpose of evaluating the probability of mTOR targeted therapy for bladder cancer. Methods After

In vivo Pharm acodynam ic Effect of Thiam phenicol in Serum of Carassius auratus on Aerom onas hydrophila

This study was to investigate the in vitro pharmacodynamic effect of thiamphenicol( TAP) in serum of Carassius auratus on Aeromonas hydrophila. By combining the in vivo pharmacokinetics and in vitro pharmacodynamics,the pharmacodynamic effect of TAP on Aeromonas hydrophila was studied,and the data were processed and analyzed by software Excel 2007,Kinetica3P97 and Kinetica4. 4. The results showed that oral administration of singly 30 mg /kg TAP assumed a rapid assimilation-quickly peaking-slowly dispelling trend in Carassius auratus. The related parameters were measured as follows: time of peaked plasma concentration of TAP( Tpeak) of 1.5 h,peak concentration( Cmax) of 37.172 μg/mL and absorption rate( ka) of 1.523 h,half-life period T1/2( ka) of 0.455 h,lag time( TL)of 0. 02 h,elimination half life T1/2( ke) of 16.712 h. The half maximal effective concentration( EC50) was 14.28 h. The PK-PD parameters were 32.41 h in AUC0- 24/ MICserumand 23. 23 in Cmax/MICserum. Employing an inhibitory Sigmoid Emax model,the administration dosage of TAP for preventing Aeromonas hydrophila-caused bacterial septicemia was 8. 61- 46. 20 mg /kg in clinical application. Based on these,we proposed the optimal administration route for preventing and controlling the Aeromonas hydrophila-caused bacterial septicemia: delivering TAP at the ratio of 46. 20 mg /kg on diseased Carassius auratus by mixing with baits or oral administration,followed by delivering with baits at ratio of 8. 61 mg /kg for preventing the Aeromonas hydrophila-caused bacterial septicemia. The results provided references for applying thiamphenicol for preventing and controlling the bacterial septicemia in aquatic livestock.

Arsenic Induced Inhibition of δ-aminolevulinate DehydrataseActivity in Rat Blood and its Response To Meso2,3-dimercaptosuccinic Acid andMonoisoamyl DMSA

Objective The objective of this study was to investigate arsenic induced changes in blood 8-aminolevulinic acid dehydratase (ALAD) after in vitro and in vivo exposure to this element and its response to co-administration of meso 2,3-dimercaptosuccinic acid (DMSA) and monoisoamyl DMSA (MiADMSA) either individually or in combination. Methods Rat whole blood was exposed to varying concentrations (0.1, 0.2 and 0.5 mmol/L) of arsenic (III) or arsenic (V), to assess their effects on blood ALAD activity. Varying concentrations of MiADMSA and DMSA (0.1,0.5 and 1.0 mmol/L) were also tried in combination to determine its ability to mask the effect of arsenic induced (0.5 mmol/L) inhibition of blood ALAD in vitro. In vitro and in vivo experiments were also conducted to determine the effects of DMSA and MiADMSA either individually or in combination with arsenic, on blood ALAD activity and blood arsenic concentration. Results In vitro experiments showed significant inhibition of the enzyme activity when 0.1-0.5 mmol/L of arsenic (III and V) was used. Treatment with MiADMSA increased ALAD activity when blood was incubated at the concentration of 0.1 mmol/L arsenic (III) and 0.1 mmol/L MiADMSA. No effect of 0.1 mmol/L MiADMSA on ALAD activity was noticed when the arsenic concentration was increased to 0.2 and 0.5 mmol/L. Similarly, MiADMSA at a lower concentration (0.1 mmol/L) was partially effective in the turnover of ALAD activity against 0.5 mmol/L arsenic (III), but at two higher concentrations (0.5 and 1.0 mmol/L) a complete restoration of ALAD activity was observed. DMSA at all the three concentrations (0.1,0.5 and 1.0 mmol/L) was effective in restoring ALAD activity to the normal value. Conclusions The results thus suggest that arsenic has a distinct effect on ALAD activity. Another important toxicological finding of the present study, based on in vivo experiments further suggests that combined administration of DMSA and MiADMSA could be more beneficial for reducing blood ALAD inhibition and blood arsenic concentration than the individual treatment.

Effects of the crude extract of Polygala tenuifolia Willd on human sperm in vitro

The aim of the present study is to analyze sperm membrane changes and the spermicidal effect in treatment with the crude extract from Polygala tenuifo/ia Willd （PTW） in vitro. The root of PTW was extracted in distilled water. Normal human spermatozoa were used to assess the spermicidal activity （Sander-Cramer assay） of the extract from the PTW root. The hypo-osmotic swelling （HOS） test and the eosin Y （EY） staining were used to detect the integrity of sperm membrane and vitality. The sperm chromatin dispersion （SCD） test was performed to determine sperm DNA integrity. N-9 was used as a reference standard and semen added to physiological saline was used as the control. Semen samples were donated by 42 healthy fertile men. The crude extract from the root of PTW could immobilize and kill 100% spermatozoa within 20 s in vitro at the concentrations of 20.0 and 10.0 mg/ml; at the concentration of 5.0 mg/ml, spermatozoa were immobilized in （39.5±3.2） s. In the groups of the crude extract from the root of PTW and N-9 solution the rate of the normal HOS （tails swollen） and the white head （unstained） was 0%, and the rate of the abnormal HOS （tails unswollen） and red head （stained） was 100%. Sperm DNA fragmentation showed no change in exposure to the crude extract from the root of PTW and N-9 solution. The sperm revival test did not show any spermatozoa that recovered their motilities. The rapid spermicidal activity of the crude extract from the root of PTW in vitro may occur by the disruption of the sperm membrane integrity.

ANTIVIRAL EFFECTS ON MOUSE LEUKEMIA VIRUS REPLICATION BY OLIGODEOXYNUCLEOTIDES IN VITRO AND IN VIVO

Oligodeoxynucleotides (Oligomers) including modified and unmodified, homo-and heterooligomers were tested for their ability to inhibit mouse SRS leukemia virus (SRSV)-induced proliferation of cells, colony formation, syncytium formation and reverse transcriptase (RT) activity in vitro. Phosphorothioate analogs complementary to Mo-MuLV sequences, as well as noncomplementary homooligomers, were found to be active. Unmodified homooligomer (dC14) also showed inhibition of growth of ascitic lymphoma carrying SRS virus. Our study suggests that different classes of oligonucleotides may inhibit SRSV replication with different mechanisms.

PLGA nanoparticle encapsulated paclitaxol for sustained release and its anticancer effect for HeLa cells in vitro

Poly (D,L-lactide-co-glycolide) (PLGA) is a biodegradable and biocompatible polymer material for drug deliver system. The aim of this study is to synthesize drug-loaded

Com bined effect of alphafetoprotein antisense oligodeoxynucleotidesand 5-fluorouracil on human hepato ma cell growth

Objective To investigate the effect of alpha fetoprotein (AFP) antisense phosphorothioate oligodeoxynucleotides in combination with 5 fluorouracil (5 FU) on the growth of BEL 7404 human hepatoma cells in vitro Methods Phosphorothioate oligodeoxynucleotides were synthesized by β cyanoethyl phosphoramidite chemistry AFP gene expression in human hepatoma cells was determined by avidin biotin peroxidase complex (ABC) immunocytochemical method Cell growth in the presence or absence of experimental agents was measured using 3 (4, 5 dimethylthiazol 2 yl) 2, 5 dipheny ltetrazolium (MTT) microculture tetrazolium assay Results AFP antisense oligomers markedly suppressed the growth of BEL 7404 human hepatoma cells in vitro by sequence specific blocking of the AFP gene expression in the cells (P<0 05) 5 FU also inhibited the hepatoma cell growth in a dose dependent manner when used alone (P<0 05) The combined treatment with AFP antisense oligomers and 5 FU showed significantly enhanced hepatoma cell growth inhibition than either AFP antisense or 5 FU treated cells alone (P<0 05) Conclusion Combined use of AFP antisense oligomers and 5 FU could more effectively inhibit the growth of BEL 7404 human hepatoma cells in vitro

Natural products and food components with anti-Helicobacter pylori activities

The bacterial pathogen Helicobacter pylori (H. pylori) colonizes in over half of the world&#x02019;s population. H. pylori that establishes life-long infection in the stomach is definitely associated with gastro-duodenal diseases and a wide variety of non-gastrointestinal tract conditions such as immune thrombocytopenia. Triple therapy which consists of a proton pump inhibitor and combinations of two antibiotics (amoxicillin, clarithromycin or amoxicillin, metronidazol) is commonly used for H. pylori eradication. Recently, the occurrence of drug-resistant H. pylori and the adverse effect of antibiotics have severely weakened eradication therapy. Generally antibiotics induce the disturbance of human gastrointestinal microflora. Furthermore, there are inappropriate cases of triple therapy such as allergy to antibiotics, severe complications (liver and/or kidney dysfunction), the aged and people who reject the triple therapy. These prompt us to seek alterative agents instead of antibiotics and to develop more effective and safe therapy with these agents. The combination of these agents actually may result in lower a dose of antibiotics. There are many reports world-wide that non-antibiotic substances from natural products potentially have an anti-H. pylori agent. We briefly review the constituents derived from nature that fight against H. pylori in the literature with our studies.

Epigenetic and Transcriptional Modulator Potential of Epigallocatechin-3-gallate and Genistein on Fetal Hemoglobin Reactivators Genes

Background:𝛽-hemoglobinopathies are one of the most common recessive genetic diseases worldwide,with limited treatments available,particularly in developed countries where the prevalence is higher.Pharmacological reactivation of Fetal Hemoglobin(HbF)is a promising therapeutic strategy.However,approximately 25%of the patients do not respond to Hydroxyurea(HU),the first and most commonly used HbF inducing agent approved by the FDA.Objective:Here,we performed an in vitro assessment of transcriptional effects induced by natural bioactive compounds,namely Epigallocatechin-3-gallate(EGCG)and genistein(GN)in globin genes(HBA1,HBB,HBG1 and HBG2)in HbF regulators/silencer genes(KLF1,BCL11A,MYB and BGLT3)and in epigenetic regulator genes(DNMT1,DNMT3A,DNMT3B,HDAC1,HDAC2,HDAC3 and HDAC8).Moreover,we evaluated EGCG’s in vivo effects in hematological parameters of healthy volunteers.Methods:K562 cells were exposed for 72 and 96 h to GN and EGCG at 100,250 and 500 ng/mL.Cell proliferation and viability were measured,and transcriptional levels were evaluated by qRT-PCR.For in vivo assay,complete blood count was determined by flow cytometry and HbF level was determined through HPLC in 30 healthy individuals before and after 225 mg EGCG ingestion per day during a 90-day period.Results:Both compounds impact cellular metabolism and proliferation with no cytotoxic effects.Divergent GN and EGCG effects in globin and BGLT3 expression levels suggest the involvement of divergent signaling pathways.As for the epigenetic potential,EGCG particularly affects HDAC2 and HDAC8 transcription,whereas GN signifi-cantly affects expression patterns of methylation and acetylation modulators.HU appears to have time divergent effects,with greater impact in methylation at 72 h(overregulates DNTM3A)while affecting acetylation mostly at 96 h(downregulates HDAC1 and HDAC8).Additionally,in vivo,EGCG demonstrated a modulator effect in hematopoiesis and HbF induction.Conclusion:Our results advocate EGCG and GN with HbF pharmacological reactivation potential and sustain further research as new alternative approaches for𝛽-hemoglobinopathies therapies.