This study evaluated the bioactive potential of heparin like polysaccharides extracted from the gladius of Sepioteuthis lessoniana (big-fin reef squid), focusing on their antibacterial, antioxidant, and anticoagulant activities. The extracted heparin-like polysaccharides were characterized using Fourier Transform Infrared Spectroscopy (FTIR), Field Emission Scanning Electron Microscopy (FESEM), and X-ray Diffraction (XRD) to determine their structural properties. Antibacterial efficacy was tested using the disc diffusion method against selected bacterial pathogens, while antioxidant activity was assessed through DPPH radical scavenging, superoxide anion scavenging, and ferrous ion chelation assays. Anticoagulant potential was evaluated using Prothrombin Time (PT) and Activated Partial Thromboplastin Time (APTT) measurements. The heparin-like polysaccharides exhibited strong antibacterial effects, with distinct inhibition zones against the tested strains. Antioxidant assays revealed significant radical scavenging and metal chelation activities, highlighting their oxidative stress mitigation potential. Moreover, the heparin-like polysaccharides prolonged PT and APTT, indicating interference with the coagulation cascade. Overall, heparin-like polysaccharides from S. lessoniana gladius demonstrate promising multifunctional bioactivities suitable for biomedical applications. However, further studies are recommended to elucidate their mechanisms of action, evaluate scalability for pharmaceutical development, and ensure long-term biocompatibility and clinical safety.

Heparosan, a glycosaminoglycan (GAG) derived from bacterial capsular polysaccharide (CPS), shares a similar backbone with both clinically utilized heparin and heparan sulfate (HS). This facilitates its transformation into heparin/HS via chemoenzymatic strategies. Currently, the purification process of heparosan, followed by N-deacetylation and N-sulfation, requires executing several complex steps prone to the unintended loss of polysaccharides and environmental risks. In this study, heparosan extracted from fermentation broth underwent immediate N-deacetylation and N-sulfation before DEAE chromatography purification. The results showed that the recovery of N-deacetylated heparosan increased from less than 40% to 93.6% with negligible contaminants. After sulfation, the overall recovery yield of N-sulfo heparosan was 76.0%. The number average molecular weight (Mn) and weight average molecular weight (MW) of N-sulfo heparosan were ascertained to be 5.2 and 10.7, respectively, with a polydispersity index (PDI) value of 2.1. The assessment of elemental composition revealed that the efficiency of N-sulfation was 84%, which aligns with that of commercial heparin. The strategy delineated in this investigation avoids the substantial loss of N-deacetylated polysaccharides resulting from the complex procedures. Furthermore, the study avoids using harmful organic solvents in preparing heparosan, thereby promoting the in vitro green synthesis of heparin-like polysaccharides and analogous pharmaceutical compounds.

Dextran sulfate sodium (DSS) is a polyanionic derivative of dextran, synthesized through the esterification of dextran with chlorosulfonic acid. DSS is a heparin-like polysaccharide; excessive use of DSS can lead to several adverse effects, including joint stiffness, pain, and hair loss. Therefore, developing a rapid and accurate method for the detection of DSS is essential for clinical diagnostics and treatment. Recently, the design of advanced photo-functional materials based on ionic liquids (ILs) has attracted significant attention due to their unique physicochemical properties, such as their ionic nature, low melting points, and non-volatile behaviour, all while maintaining intrinsic photoluminescent functionality. In this study, we have strategically developed a fluorescent ionic liquid (FLIL) through a simple ionic exchange reaction. The resulting material exhibits high sensitivity and selectivity towards DSS in both solid and liquid phases. The pure FLIL displays orange colour photoluminescence under a 365 nm portable UV lamp. When converted into water-suspended nanoparticles, nFLIL, the system shows a remarkable transition from non-fluorescent to green fluorescent upon the stepwise addition of DSS. The probe demonstrates excellent selectivity toward DSS with a limit of detection and quantification in the nanomolar range. In addition, a portable and user-friendly paper-based strip kit was developed, highlighting its potential for practical applicability of DSS detection in solid-phase samples. This work introduces an innovative approach for the development of DSS-sensitive, IL-based materials, offering superior properties over conventional systems for potential use in forensic and environmental monitoring.

Ischemic wounds are chronic wounds with poor blood supply that delays wound reconstruction. To accelerate wound healing and promote angiogenesis, adipose-derived stem cells (ADSCs) are ideal seed cells for stem cell–based therapies. Nevertheless, providing a favorable environment for cell proliferation and metabolism poses a substantial challenge. A highly sulfated heparin-like polysaccharide 2-N, 6-O-sulfated chitosan (26SCS)-doped poly(lactic-co-glycolic acid) scaffold (S-PLGA) can be used due to their biocompatibility, mechanical properties, and coagent 26SCS high affinity for growth factors. In this study, a nano-scaffold system, constructed from ADSCs seeded on electrospun fibers of modified PLGA, was designed to promote ischemic wound healing. The S-PLGA nanofiber membrane loaded with adipose stem cells ADSCs@S-PLGA was prepared by a co-culture in vitro, and the adhesion and compatibility of cells on the nano-scaffolds were explored. Scanning electron microscopy was used to observe the growth state and morphological changes of ADSCs after co-culture with PLGA electrospun fibers. The proliferation and apoptosis after co-culture were detected using a Cell Counting Kit-8 kit and flow cytometry, respectively. An ischemic wound model was then established, and we further studied the ability of ADSCs@S-PLGA to promote wound healing and angiogenesis. We successfully established ischemic wounds on the backs of rats and demonstrated that electrospun fibers combined with the biological effects of adipose stem cells effectively promoted wound healing and the growth of microvessels around the ischemic wounds. Phased research results can provide a theoretical and experimental basis for a new method for promoting clinical ischemic wound healing.

Chemical and biological differences between original and mimetic pentosan polysulfates

Pentosan polysulfate sodium (PPS) is a heparin-like polysaccharide that is applied as atherapeutic treatment for osteoarthritis (OA) in animals. This study investigated theefficacy of different molecular weights PPS (1,500–7,000 Da) on the phenotype regulatoryand chondrogenic properties of canine articular chondrocytes. The cytotoxicity of PPS onchondrocytes was assessed using flow cytometry and3-(4,5-dimehylthiazolyl-2)-2,5-diphenyltetrazolium bromide assay. After 72 hr of exposure,PPS did not induce chondrocyte apoptosis, regardless of molecular weight. In addition,chondrogenic properties were determined according to the mRNA and protein levels inmicromass-cultured chondrocytes. Quantitative polymerase chain reaction analysis confirmedthat PPS promotes a chondrogenic phenotype in chondrocytes in a molecular weight-dependentmanner, with significant upregulation of collagen type II alpha 1 chain,aggrecan, and SRY-box transcription factor 9(SOX9) mRNA levels relative to those in the control. However, thecollagen type I alpha 2 chain mRNA level simultaneously increased after7,000 Da PPS treatment. PPS exposure also increased collagen type II and SOX9 proteinproduction in a molecular weight-dependent manner and inhibited Akt phosphorylation inchondrocytes. Alcian blue staining indicated that PPS treatment enhanced proteoglycandeposition in micromass cultures, with stronger effects observed in 5,000 and 7,000 Dagroups. Overall, these results indicate that PPS exerts protective effects on thechondrocyte phenotype and may represent a potential therapeutic target for OA treatment.Increasing the molecular weight of PPS could enhance these anabolic effects.

Preparation of fully bio-based multilayers composed of heparin-like carboxymethylcellulose sodium and chitosan to functionalize poly (l-lactic acid) film for cardiovascular implant applications

Biodegradable reduction-responsive polymeric micelles for enhanced delivery of melphalan to retinoblastoma cells

Emerging evidence suggests that successful healing of bone substitutes depends on the osteogenesis-angiogenesis interplay. Bone morphogenetic protein-2 (BMP-2) and vascular endothelial growth factor (VEGF) have been identified as key regulators of osteogenesis and angiogenesis during bone regeneration. While the importance of growth factors is now widely accepted, the impact and mechanisms of different releasing sequences on bone repair have not been fully understood. Here, a composite vehicle (Gel/PMs), constructed with hydrogels and microspheres, was developed, which is capable of achieving two distinct releasing modes: BMP-2 first release followed by VEGF release (B/V) and VEGF first release followed by BMP-2 release (V/B). In our results, the B/V mode exhibited more extensive vascular network formation by up-regulating angiogenic genes during the bone remolding, thus facilitating rapid bone transformation which was confirmed by radiography. Further histological and immune-staining analysis revealed that fast release of BMP-2 made for rapidly initiating osteogenesis, while later VEGF release promoted persistent angiogenesis and mature bone formation. Moreover, interest arises from the introduction of 2-N,6-O-sulfated chitosan (SCS), a sulfonated heparin-like polysaccharide. It has synergistic effects with both BMP-2 and VEGF, which can further accelerate bone healing by efficiently improving osteogenesis and angiogenesis. The results demonstrated that disparate releasing sequence of growth factors might influence regenerative efficiency. Such a strategy may provide insights toward designing bioactive materials and give promising application in tissue regeneration.

Efficacy of heparinoid PSS in treating cardiovascular diseases and beyond-A review of 31 years clinical experiences in China.

The inhibitory effects and mechanisms of 3,6-O-sulfated chitosan against human papillomavirus infection

Pathogenic bacterial infection has been becoming a global threat toward people's health, especially the massive usage of antibiotics due to the lack of antibacterial agents with less side effects. Developing new nanoagents to fight pathogenic bacteria has provided enormous new possibilities in the treatment of bacterial infections, such as graphene-based two-dimensional (2D) antibacterial nanoagents with different bacterial inhibition capabilities; however, mussel-inspired design of near-infrared (NIR)-responsive and biocompatible Ag-graphene nanoagents possessing efficient and versatile bacterial disinfection activities have rarely been reported. In this study, we developed a new kind of antibacterial nanoagent, dopamine-conjugated polysaccharide sulfate-anchored and -protected Ag-graphene (Ag@G-sodium alginate sulfate ((SAS)) nanocomposite, to combat bacterial infection and contamination in different application fields. Ag@G-SAS exhibited robust antibacterial activity toward both Escherichia coli and Staphylococcus aureus; notably, the nanoagent can significantly inhibit S. aureus infection on wounded pig skin without or with NIR laser. Besides wound disinfection, the 2D Ag@G-SAS can also serve as a good layer-by-layer (LbL) building block for the construction of self-sterilizing coatings on biomedical devices. All of the results verified that the LbL-assembled Ag@G-SAS coating exhibited favorable bactericidal activity, extraordinary blood compatibilities, and good promotion ability for cell proliferation. Owing to the shielding effects of heparin-like polysaccharide sulfates, the Ag@G-SAS nanoagent showed limited cytotoxicity toward mammalian cells. Combining all of the advantages mentioned above, it is believed that the proposed Ag@G-SAS nanoagent and its LbL-assembled coatings may have versatile application potentials to avoid bacterial contaminations in different fields, such as wounded skin, disinfection of biomedical implants and devices, and food packaging sterilization.

Inductive influence of heparin-like polysaccharide on the keratinocyte differentiation

TGF-β regulates several steps in cancer metastasis, including the establishment of bone metastatic lesions. TGF-β is released from bone during osteoclastic bone resorption and it stimulates breast cancer cells to produce osteolytic factors such as interleukin 11 (IL-11). We conducted a cell-based siRNA screen and identified heparan sulfate 6-O-sulfotransferase 2 (HS6ST2) as a critical gene for TGF-β-induced IL-11 production in highly bone metastatic MDA-MB-231(SA) breast cancer cells. HS6ST2 attaches sulfate groups to glucosamine residues in heparan sulfate glycosaminoglycans. We subsequently showed how heparin and a high-molecular-weight Escherichia coli K5-derived heparin-like polysaccharide (K5-NSOS) inhibited TGF-β-induced IL-11 production in MDA-MB-231(SA) cells. In addition, K5-NSOS inhibited bone resorption activity of human osteoclasts in vitro. We evaluated the therapeutic potential of K5-NSOS and fragmin in a mouse model of breast cancer bone metastasis. MDA-MB-231(SA) cells were inoculated into the left cardiac ventricle of athymic nude mice which were treated with fragmin, K5-NSOS, or vehicle once a day for four weeks. Both heparin-like glycosaminoglycans inhibited weight reduction, decreased osteolytic lesion area, and reduced tumor burden in bone. In conclusion, our data imply novel mechanisms involved in TGF-β induction and support the critical role of heparan sulfate glycosaminoglycans in cancer metastasis as well as indicate that K5-NSOS is a potential antimetastatic and antiresorptive agent for cancer therapy. This study illustrates the potential to translate in vitro siRNA screening results toward in vivo therapeutic concepts.

The use of SAX-HPLC–CD as a heparin screening strategy

Abstract Transforming growth factor β (TGF-β) is a key regulator of several steps in cancer metastasis, including the establishment of bone metastatic lesions. TGF-β is released from bone during osteoclastic bone resorption and it stimulates breast cancer cells to produce osteolytic factors such as interleukin 11 (IL-11). We conducted a cell-based siRNA screen and identified heparan sulfate 6-O-sulfotransferase 2 (HS6ST2) as a critical gene for TGF-β-induced IL-11 production in highly bone metastatic MDA-MB-231(SA) breast cancer cells. HS6ST2 attaches sulfate groups to glucosamine residues in heparin sulfate which is a member of the glycosaminoglycan family of carbohydrates. Our subsequent studies with highly sulfated heparin-like glycosaminoglycans (HLGAGs) with low anti-coagulant activity indicated that a high-molecular-weight E. coli K5-derived heparin-like polysaccharide (K5-NSOS) inhibited TGF-β-induced IL-11 production in MDA-MB-231(SA) cells. We then evaluated the therapeutic potential of K5-NSOS and a low-molecular-weight synthetic heparin (fragmin) in a mouse model of breast cancer bone metastasis. MDA-MB-231(SA) cells were inoculated into the left cardiac ventricle of athymic nude mice which were then administered with K5-NSOS, fragmin, or vehicle once daily for 4 weeks. Both HLGAGs increased body weight, decreased osteolytic lesion area, and reduced tumor burden in bone. Our in vitro studies showed that both K5-NSOS and fragmin inhibited adhesion of breast cancer cells to ICAM-1, and K5-NSOS also inhibited adhesion to E- and P-selectin. Furthermore, K5-NSOS inhibited bone resorption activity of human osteoclasts in vitro. Our data support and further clarify the critical role of heparan sulfate glycosaminoglycans in breast cancer metastasis and indicate that K5-NSOS is a potential antimetastatic and antiresorptive agent with low anticoagulant activity that could be further optimized as an anti-tumor agent. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 845. doi:10.1158/1538-7445.AM2011-845

Percutaneous coronary interventions play a major role in the management of patients affected by coronary artery diseases. However, their efficiency is impaired by restenosis, defined as a reduction of the vessel lumen, occurring a few months after the procedure. A low-molecular-weight fraction of fucoidan, a vegetal heparin-like sulphated polysaccharide, was recently shown to greatly reduce in-stent restenosis after angioplasty in rabbits. To better understand the in vivo anti-restenotic effects of this polymer, we used fractions of fucoidan and compared to heparin and dextran of different sizes. We carried out in vitro growth inhibition experiments on vascular smooth muscle cells, performed an in vivo pharmacokinetic study, and locally delivered fluorescently-labeled polysaccharides in rabbit iliac arteries after angioplasty with a non-occlusive catheter. The results indicated that (i) preparation of well-characterized fractions from natural fucoidan is compulsory for in vitro and in vivo studies, (ii) antiproliferative activity of sulphated polysaccharides on cultured smooth muscle cells is not a major predictive factor for the reduction of restenosis in vivo and (iii) pharmacokinetic parameters and binding of low-molecular-weight fucoidan on angioplasty-induced injured vascular walls are important local and general factors controlling its mechanisms of action.

Heparin-like polysaccharides possess the capacity to inhibit cancer cell proliferation, angiogenesis, heparanase-mediated cancer cell invasion, and cancer cell adhesion to vascular endothelia via adhesion receptors, such as selectins. The clinical applicability of the antitumor effect of such polysaccharides, however, is compromised by their anticoagulant activity. We have compared the potential of chemically O-sulfated and N,O-sulfated bacterial polysaccharide (capsular polysaccharide from E. COLI K5 [K5PS]) species to inhibit metastasis of mouse B16-BL6 melanoma cells and human MDA-MB-231 breast cancer cells in two in vivo models. We demonstrate that in both settings, O-sulfated K5PS was a potent inhibitor of metastasis. Reducing the molecular weight of the polysaccharide, however, resulted in lower antimetastatic capacity. Furthermore, we show that O-sulfated K5PS efficiently inhibited the invasion of B16-BL6 cells through Matrigel and also inhibited the in vitro activity of heparanase. Moreover, treatment with O-sulfated K5PS lowered the ability of B16-BL6 cells to adhere to endothelial cells, intercellular adhesion molecule-1, and P-selectin, but not to E-selectin. Importantly, O-sulfated K5PSs were largely devoid of anticoagulant activity. These findings indicate that O-sulfated K5PS polysaccharide should be considered as a potential antimetastatic agent.

Pentosan polysulfate (PPS) is a heparin-like polysaccharide that can affect the binding interactions of fibroblast growth factor (FGF-2) with its high-affinity receptors. Patients with angiogenic tumors frequently show high levels of FGF-2 in the circulation. Since FGF-2 is a heparin-binding angiogenic growth factor, PPS has been used successfully to block its activity in patients with angiogenic tumors. However, because of its heparin-like activity, the major toxic effect of PPS is the development of bleeding disorders. The role that circulating FGF-2 plays in the pathogenesis of bleeding disorders in patients treated with PPS is currently unknown. Here we hypothesized that FGF-2 might play a physiological role in the pathogenesis of intestinal bleeding induced by PPS. This hypothesis is supported by previous studies showing that PPS is accumulated in the intestine and that circulating FGF-2 specifically binds to and modulates the angiogenic activity of intestinal submucosal endothelial cells. We used recombinant adenoviral vectors carrying a secreted form of FGF-2 and LacZ control vectors to determine whether high levels of circulating FGF-2 facilitate the development of intestinal bleeding disorders in FVB/N and C57BL/6J mice treated with PPS. We found that PPS, acting together with FGF-2, induced structural changes in intestinal vessels leading to the development of lethal intestinal hemorrhages. These findings might have wider clinical implications for the systemic use of PPS and other heparinoids in the treatment of patients with angiogenic diseases associated with high levels of circulating FGF-2.

Heparan sulfate carries a wide range of biological activities, regulating blood coagulation, cell differentiation, and inflammatory responses. The sulfation patterns of the polysaccharide are essential for the biological activities. In this study, we report an enzymatic method for the sulfation of multimilligram amounts of heparan sulfate with specific functions using immobilized sulfotransferases combined with a 3'-phosphoadenosine 5'-phosphosulfate regeneration system. By selecting appropriate enzymatic modification steps, an inactive precursor has been converted to the heparan sulfate having three distinct biological activities, associated with binding to antithrombin, fibroblast growth factor-2, and herpes simplex virus envelope glycoprotein D. Because the recombinant sulfotransferases are expressed in bacteria, and the method uses a low cost sulfo donor, it can be readily utilized to synthesize large quantities of anticoagulant heparin drug or other biologically active heparan sulfates.